AIDS Indicator Survey

HIV Testing Manual

Macro International Inc.

Calverton, Maryland

April 2007
The AIDS Indicator Survey was developed as part of the MEASURE DHS project. MEASURE DHS
assists countries worldwide with the collection and use of data to monitor and evaluate population, health
and nutrition programs. Funded by the U.S. Agency for International Development (USAID), MEASURE
DHS is implemented by Macro International Inc. in Calverton, Maryland.

Information about the MEASURE DHS project can be obtained by contacting Macro International Inc.,
11785 Beltsville Drive, Suite 300, Calverton, MD, 20705, USA. Telephone: 301-572-0200; Fax: 301-
572-0999; E-mail: [email protected]; Internet: http://www.measuredhs.com.

Suggested citation:
Macro International. 2007. AIDS Indicator Survey: HIV Testing Manual MEASURE DHS Calverton,
Maryland, USA.: Macro International.
Contents

SECTION 1: INTRODUCTION ............................................................................................................... 1

1.1 General Background .................................................................................................................... 1

1.2 HIV Testing ................................................................................................................................. 1
1.3 Training of Interviewers on HIV Testing .................................................................................... 2
1.4 Supervision of Interviewers During Blood Collection ................................................................ 3
1.5 Organization of the Training Manual .......................................................................................... 3

SECTION 2: MATERIALS AND EQUIPMENT FOR TESTING........................................................ 5

2.1 Materials and Supplies for Performing Finger Prick................................................................... 5

2.2 Materials and Supplies for HIV Testing...................................................................................... 6

SECTION 3: COMPLETING QUESTIONNAIRES AND OTHER TESTING DOCUMENTS........ 8

3.1 Identifying Eligible Respondents ................................................................................................ 8

3.2 Obtaining Informed Consent for the Testing............................................................................... 9
3.3 Recording Information on the Test Schedules........................................................................... 11
3.4 Providing VCT Information ...................................................................................................... 12

SECTION 4: PROCEDURES FOR COLLECTING CAPILLARY BLOOD DROP SAMPLES .... 16

4.1 Steps in Obtaining Capillary Blood from the Finger................................................................. 16

SECTION 5: HIV TESTING FOR ADULTS AND YOUTHS ............................................................. 19

5.1 Collecting Blood for HIV Testing ............................................................................................. 19

5.2 Providing Referrals for HIV VCT Material............................................................................... 21
5.3 Storing and Transferring the Dried Blood Spots ....................................................................... 21

SECTION 6: PRECAUTIONS TO OBSERVE WHEN COLLECTING BLOOD SAMPLES ........ 24

6.1 General Precautions when Collecting Blood............................................................................. 24

6.2 Rules for Collecting Specimens for HIV Testing...................................................................... 24

SECTION 7: BIOHAZARDOUS WASTE DISPOSAL ........................................................................ 26

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SECTION 1: INTRODUCTION

1.1 General Background

This manual discusses in detail the techniques and procedures for collecting, handling and packaging
blood samples for HIV testing during the AIDS Indicator Survey (AIS). In order to collect the blood
samples for HIV testing, the interviewers for the AIS will act as both interviewers and as medical staff
who will also be responsible for taking the blood samples from eligible women and men. In additional to
their duties as interviewers, they will be responsible for:

• locating all eligible women and men for HIV testing;

• informing eligible women and men about the test and asking for consent;

• collecting and labeling blood samples;

• filling the necessary forms;

• leaving a brochure with HIV information;

• telling respondents where they can go for HIV testing if they want to know their HIV status;

• giving them a voucher for free testing.

The AIDS Indicator Survey will collect information on demographic characteristics, reproduction,
marriage and sexual activity, HIV/AIDS and sexually transmitted infections (STIs) through individual
interviews with a representative sample of women and men in the reproductive ages. In addition to
individual interviews, blood samples will be collected in the survey households from men and women age
15-49 years for HIV testing.

1.2 HIV Testing

Acquired immunodeficiency syndrome (AIDS) is a condition caused by human immunodeficiency virus
(HIV). It is a disorder of the immune system in which the body’s normal defenses against infection break
down, leaving it vulnerable to a host of life threatening infections, including unusual malignancies.

HIV testing in the AIS will involve the collection of dried blood spots (DBS) samples on a filter paper
card. This is performed using capillary blood from a finger stick. After the DBS samples and
questionnaires have been transported to the central office for logging and checking, the DBS samples will
be transported to the laboratory where the testing will be conducted. The testing will be anonymous, i.e.,
personal identifiers of individual respondents will not be retained with the DBS samples. Because the
testing is anonymous, respondents will not receive the results of the testing. However, they will be offered
a coupon to obtain free HIV counseling and testing at a collaborating VCT center(s) near their
community.

Testing of Men and Women

Identify all men and women from the household schedule who are eligible for testing. Women and men
age 15-49 years will be eligible for HIV testing;

Check the age of the respondent recorded in question 103. Then go to the HIV Test section and record
whether the respondent is age 15-17 or age 18-49 for Question 701. If the respondent is age 15-17, record

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whether the respondent has ever been married in Question 702. You will do this because if the
respondent is age 15-17 and has never been married, consent for testing will have to be obtained from the
respondent’s parent or responsible adult;

If the respondent is between ages 15-17 and has never been married, record name and line number of the
parent or adult responsible for the youth in Question 703;

If the respondent is age 15-17 and has never been married, seek consent for HIV testing from the
parent/responsible adult before seeking consent directly from the respondent. If the eligible respondent is
age 18 or over or has ever been married, seek direct consent of the respondent for HIV testing. Record
outcome of the consent request in Question 704. If the parent refuses to consent, circle 2. If the
respondent refuses to consent, circle 3. If consent is granted, circle 1. The signature in Question 704 will
be your signature, confirming that you read the statement to the responsible adult;

After obtaining the consent, identify the next available complete set (row) of bar code labels. Paste the
first bar code label in Question 705, the second bar code label on a new filter paper card, and the third bar
code label on the transmittal sheet;

Obtain a capillary blood sample from the finger prick. Use enough blood drops to fill as many of the
preprinted circles on the filter paper card as possible for HIV antibody testing in the laboratory. At least
four of the circles must be filled.

Place the filter paper card with the blood spots in the drying rack;

Record the result for HIV blood sample collection in Question 706;

Seek consent to store part of the blood sample for future research. If the respondent is age 15-17 and has
never been married, seek consent for blood storage from the parent/responsible adult before seeking
consent directly from the respondent. If the eligible respondent is age 18 or over or has ever been married,
seek direct consent of the respondent for HIV testing. Record outcome of the consent request in Question
707. If the parent refuses to consent, circle 2. If the respondent refuses to consent, circle 3. If consent is
granted, circle 1. The signature in Question 707 will be your signature, confirming that you read the
statement to the responsible adult;

Provide an informational brochure on HIV/AIDS, a coupon for free VCT services, and a list of the nearest
participating VCT centers.

Subsequent sections in this manual will describe in detail the procedures you will follow in accomplishing
the above tasks.

1.3 Training of Interviewers on HIV Testing

Your training to conduct the HIV testing will consist of a combination of classroom training and practical
experience that will provide you with the background necessary to perform these tasks during a field
survey.

During the first phase of training, we will review with you various sections of this manual. You will learn
how to identify eligible respondents, record information relating to the testing in the household
questionnaire on special field forms, handle the technical procedures involved in specimen collection,
testing and transporting, and other related instructions.

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During the second phase of the training, there will be role-playing for HIV testing in which you will
practice by collecting blood samples from other trainees. During the third phase of the training, we will
visit a health center and practice testing on consenting clients.

During the final part of the training, you will be assigned to an AIS trainee team and will practice both
interviewing and collecting blood samples from eligible adults in some pretest areas outside of those
chosen for the AIS sample.

Before each training session, you should study this manual carefully along with the household
questionnaire, writing down any questions you may have. You are encouraged to ask questions at any
time in order to avoid mistakes during actual fieldwork. You can learn a lot from each other by asking
questions and talking about problems encountered in practice and actual situations.

Like the rest of the AIS training, throughout the blood collection training, you will be given homework
assignments for the evenings. You may be given tests to see how well you are progressing during your
formal training period. At the end of the training course, your overall performance during the training will
be assessed and we will select those of you who have performed the best to work in the main survey.

1.4 Supervision of Interviewers During Blood Collection

Training is a continuous process. Observation and supervision throughout the fieldwork are a part of the
training and data collection process. Your team supervisor and the AIS survey coordinators will play very
important roles in continuing your training and in ensuring the quality of data. They will:

• observe some of your fieldwork activities to ensure that you are conducting yourself
professionally, asking the questions in the right manner, and following the survey protocol
correctly;

• spot check some of the eligible respondents selected for testing to be sure that you collected the
blood specimen from the correct households and respondents;

• consolidate the field record forms, samples, and both the household and individual
questionnaires;

• meet with each member of the team on a regular basis to discuss performance and give out future
work assignments;

• help to solve any problems that you might have with finding the assigned households,
understanding the concepts in the questionnaire or dealing with difficult respondents.

The survey director may release from service any field staff member who is not performing at the level
necessary to produce the high quality data required to make the AIS a success.

1.5 Organization of the Training Manual

The remaining sections of this manual are organized to assist you in learning how to conduct these tasks.
They include:

Section 2: Introduction to the equipment and supplies for the testing

Section 3: Recording information on the testing
Section 4: General procedures to follow in obtaining a capillary blood sample

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 Section 5: Specific procedures to follow in collecting blood samples for HIV
Section 6: Precautions to follow in collecting samples
Section 7: Disposing of biohazardous waste
These sections should be studied carefully during the training and used as a reference in the field when
you have questions about how you should proceed in conducting the various tasks involved in testing.

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SECTION 2: MATERIALS AND EQUIPMENT FOR TESTING

This section of the manual introduces the various materials and supplies which you will use for HIV
testing. More detail on the use of these items is presented in Sections 4-6, which describe how the testing
will be performed.

2.1 Materials and Supplies for Performing Finger Prick

The capillary blood drops used for HIV testing will be drawn from a finger. The following supplies and
materials (Figure 2.1) will be used in performing the finger prick:

• Non-powdered disposable latex gloves are used to reduce the risk of blood-borne diseases.
Gloves must be worn by the interviewer and by anyone else who may assist with the blood
collection.

• Alcohol preps are used for cleaning the skin prior to puncturing the finger.

• Sterile gauze pads are used to wipe away the first drops of blood to stimulate a spontaneous
capillary blood flow.

Figure 2.1 Supplies for performing a finger prick

• Adhesive bandages (Band-aids). After the blood collection, an adhesive bandage has to be
applied on the puncture site to avoid infection.

• Retractable lancets. The lancet is an automated, disposable incision device used to puncture the
fingertip. The device is specially shaped to fit easily on the skin surface, thus minimizing skin
indentation; the angle of the blade is set for maximum blood flow. When the trigger is pressed, a
surgical blade quickly protrudes from the device and then automatically retracts and the action of
the blade is so fast it cannot be seen.

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2.2 Materials and Supplies for HIV Testing
The following special equipment and supplies are used in the collection, storage and transport of the dried
blood spot (DBS) samples.

• Desiccants (drying agents) (Figure 2.2). Desiccant

packets, which absorb moisture from the air, are used to
keep the materials used for collecting the DBS samples as
dry as possible. Non-regeneratable desiccant packets are
transparent packets in which the granules are visible. The
granules change color as they absorb moisture. You should
change the desiccants as indicated by the humidity
indicator cards or when the packet has changed color.

• Humidity indicator cards (Figure 2.3). The non-

regeneratable desiccant packets will turn color at the point
Figure 2.2 Desiccants
of saturation. The humidity indicator card allows you to
more closely monitor the level of moisture. This will
enable you to add additional desiccant packets in
conditions of high humidity.

There are three circles on the humidity indicator card. If

the circle at the bottom of the card (labeled 30%) turns
pink, it indicates a relatively high level of humidity and is
a warning to begin to carefully monitor the humidity level.
If the middle or top circles (labeled 40% and 50%,
respectively) turns pink, you should replace the desiccant
packets in the bag with fresh packets. Replace the
humidity indicator card with a fresh card if the circles
merge.

• Filter paper card (Figure 2.4). You will use special filter
paper cards to collect the blood samples. Each of the cards Figure 2.3 Humidity Indicator Card
has five preprinted circles that contain approximately 100
µl blood when fully filled. You must fill at least four of the
circles for the HIV testing.

The preprinted circles of the filter paper cards must be

kept clean and dry at all times as water, dust, sweat from
your hands, or other environmental contaminants can
affect the HIV test results. Thus, you should use gloves at
all times when handling the filter paper cards.

• The filter paper cards come in packets of 100 cards. When

you need a new packet of filter paper cards, put on gloves
first. Open the packet and place all cards in a large zip-loc
bag. They must be stored stacked so as to avoid
Figure 2.4 Filter paper cards
compressing the filter paper. Place two non- (903 TM paper)
regeneratable desiccant packets and a humidity
indicator card in each bag before sealing it. Later, if the
humidity indicator card changes color, follow the above instructions. If you do not have humidity

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 indicator cards and the crystals in the desiccant packets change color before you have used up all
of the 100 filter paper cards, you should replace the desiccant packets with fresh packets.

• Bar code labels. Because the HIV testing is anonymous, bar code labels will be used to identify
the DBS samples. These peel-off, preprinted bar code labels are provided on special sheets. Each
row on a sheet includes a number of labels with the same bar code. A different row on the sheet
will be used for each respondent for whom a DBS sample is collected. Instructions for using the
bar code labels are included in Section 6 of this manual.

• Drying Box. After collection, the blood spot samples must be placed in a specially designed box
where they will be stored overnight to dry. The plastic drying box has a cardboard paper drying
rack that holds the sample filter paper cards in a horizontal position when the box is standing
vertically. The plastic boxes are to be used for collection and overnight drying only. They are not
intended for long-term sample storage. Pouches of non-regeneratable desiccants should be kept at
all times at the bottom of the drying box (as it stands vertically) to hasten the drying process,
particularly in high humidity environments. A humidity indicator card should be placed in the
drying box to allow you to monitor the level of humidity.

• Low gas-permeable bags (small zip-loc bags). You will be given a supply of special small zip-
loc bags to use for storing samples and supplies in the field. These bags are specially
manufactured to reduce the exposure of their contents to air and moisture. These bags are
expensive and should never be used for other purposes, like carrying food or adhesive bandages.
The bags have a sliding “zipper” that is used to close and seal the bag. Each morning, you will
transfer the DBS samples from the previous day’s fieldwork, which have dried overnight, to these
bags (See Section 6).

• Glassine paper. Thin, glossy, semi-opaque paper squares. The glassine paper is used to protect
the dried blood spots on the filter paper cards during storage.

• Large zip-loc bags. A large zip-loc bag will be provided to you for each of the AIS sample
clusters in which you will work. These large zip-loc bags will be used to hold the DBS samples
from the cluster during storage and transport.

• Plastic bags for biohazardous wastes. These are big bags that are provided to hold all the
biohazardous waste materials during the day and will be discarded appropriately (see Section 7).

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SECTION 3: COMPLETING QUESTIONNAIRES AND OTHER
TESTING DOCUMENTS

You will be responsible for the accurate recording of information that will be used to track the outcome of
the testing procedures. Documents which you will complete during the testing are:

• Blood Spot Collection page for women and men in the Individual Questionnaire.

• Blood Sample Transmittal Sheet.

• Voucher/coupon for free VCT services.

In addition, you will be provided with informational brochure on HIV/AIDS – to be left with the
households that you visit for blood collection for HIV testing.

After the fieldwork and data entry and cleaning have ended, the Blood Spot Collection page, which
includes the unique barcode assigned to the respondent’s HIV blood sample, will be removed from the
questionnaire in the central office and destroyed. This is to ensure the anonymity of the HIV test results. It
will be impossible to identify which respondent gave a particular blood sample for testing once the page
is removed and destroyed.

This section reviews the various tasks that are associated with using these various materials. The key
tasks include:

• Identifying eligible respondents.

• Obtaining informed consent for the testing.

• Recording information relevant to the outcome of the testing in the Individual Questionnaire and
other tracking documents.

• Providing informational brochures and written referrals for HIV VCT.

Specific activities that are involved in performing these tasks are described in detail below.

3.1 Identifying Eligible Respondents

For eligible women and men (age 15-49 years), information relating to HIV testing will be recorded on
the HIV Test page found in the Individual Questionnaire.

The first step in the testing process will be to identify members of the household who are eligible for HIV
testing. All women and men age 15-49 years are eligible for collection of dried blood spot samples for
later HIV testing in the laboratory. The age of the respondent has to be recorded and checked from
question 103 of the individual questionnaire.

All eligible consenting respondents should be tested after their individual interview has been
completed.

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3.2 Obtaining Informed Consent for the Testing
One of the primary tasks before blood collection is to explain the purpose of the testing to eligible
respondents and to obtain their consent before collecting any blood samples. In order to ensure that these
individuals can make an ‘informed’ decision about the testing, the AIS questionnaire includes statements
which you will read as appropriate. These ‘informed consent’ statements (Figures 3.1 and 3.2) include the
following basic elements:

1. a description of the objectives of the test

2. basic information on how the test will be conducted

3. assurances about the confidentiality of the results

4. a specific request for permission to collect the sample.

You must read the informed consent statements to each eligible respondent age 18 and over and obtain
the respondent’s consent to the testing before any blood collection is done. The approach for obtaining
consent differs slightly when the eligible respondent is a never-married youth (15-17 years old). If the
respondent is a never-married youth, you must first obtain the consent of one of the respondent’s parents,
or, in the absence of a parent, the consent of another responsible adult who is at least 18 years of age. For
these respondents, after obtaining consent from the parent/responsible adult, you must read the consent
statement to the youth. The only exception is if the youth lives alone or in a household in which there are
no adults. In such cases, the consent of the youth is sufficient.

In all cases, you must record the outcome of the consent request before performing the test on a
respondent.

The informed consent process for HIV testing for youths involves several steps. The following
information must be recorded on Blood Spot Collection/ HIV Testing Page:

• Question 701: Verify the age of the respondent.

• Question 702: Verify that the respondent has never been married. As noted above, if the
respondent is age 15-17 and not married or not living independently, check the appropriate boxes
in Questions 701 and 702. You must obtain permission from his/her parent/responsible adult
before seeking the respondent’s own consent for the testing.

• Question 703: For never-married respondents age 15-17, record the line number of the
parent/responsible adult whose consent for the testing will be requested.

Record whether both the parent/responsible adult and the respondent consent to the HIV test in Question
704. If both the respondent and the parent/responsible adult consent for the test to be conducted on the
youth, circle 1. If the parent refuses, circle 2. If the respondent refuses, circle 3, even if the respondent’s
parent consented.

You must sign your name to indicate that you read the consent statements to both the respondent and the
parent/responsible adult in the case of non-married, non-independent youths and have recorded their
response accurately.

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 Figure 3.1 Consent Statement for HIV Testing

The following are some key points to remember in obtaining consent for the testing:

1. Read the applicable consent statements to each eligible respondent exactly as they appear in the
questionnaire. When you arrive at the household and begin talking about the blood tests with an
eligible respondent, you may informally discuss many of the items included in the informed consent
statement. However, before beginning the testing with any respondent, you must still read the
informed consent statements exactly as they are worded in the questionnaire. If you feel that the
respondent may find the statements repetitive, tell him/her that you are required to formally read the
statement to ensure that respondents are given all the appropriate information.

2. The only exception that is allowable is in cases when you have already requested consent for testing
from an adult who also happens to be the parent or other responsible adult who must give consent to
test a youth in the household. In such cases, you can simply ask the parent/responsible adult: “Do I
also have your permission for (NAME OF YOUTH) to participate in the study?”.

3. Be sure that you read the informed consent statements clearly. Practice reading the consent
statements so that you become comfortable reading them in a clear, natural manner. Avoid using a
monotone when reading the statements or reading them so rapidly that they cannot be understood.

4. Never collect blood from a never-married youth (15-17 years) before obtaining the consent of
the parent/responsible adult unless the youth lives alone or in a household where there are no adult
members.

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5. Never attempt to force or coerce consent. It may take tact and patience to overcome people’s fears
about having blood collected for testing. Take your time in trying to convince respondents who are
uncertain about the testing to grant their consent. Some respondents may have questions or want to
discuss the procedures before giving consent. Patiently respond to all questions.

6. Some respondents from whom you are seeking consent may be reluctant to allow any testing without
consulting someone not present at the time of your visit (such as a woman who may want to consult
her husband before giving permission). In such cases, make an appointment to return to the household
later at an agreed upon time. If you believe it will help, ask the team supervisor to visit a household
where eligible respondents express fear or reluctance to be tested.

3.3 Recording Information on the Test Schedules

Information relevant to the outcome of the testing process must also be recorded on the Blood Spot
collection page (Figure 3.2).

• Record the final outcome of the HIV testing in Question 706. Record ‘1’ if the respondent was
successfully tested, i.e. the sample for HIV testing was successfully taken from the respondent,
‘2’ if the respondent was not present at the time you visited the house for testing, ‘3’ if the
respondent (or the parent/responsible adult for respondents age 15-17) refused consent for the
testing, Use code ‘6’ if you have technical problems or for other reasons the test could not be
performed (e.g., the respondent was drunk or otherwise incapacitated) and specify the reason.

Figure 3.2 Recording the Outcome of the Testing Process

The following summarizes the procedures for recording this information for adults and youths for the
Blood Spot Collection Page and the Blood Sample Transmittal Sheet:

To ensure that the testing process is accurately documented for each eligible respondent, it is important
that you fill in all of the relevant items in these documents in the following sequence:

If all necessary consents have been obtained for HIV testing, you will:

• Assign HIV blood sample bar code. Each respondent who consents to the HIV testing will be
assigned a unique identification number from the sheet of preprinted bar code labels you will be
provided. The labels come in sets of three across a sheet.

• Paste the first bar code label in Question 705 of the HIV Test page in the Individual
Questionnaire. Paste the second bar code label with the same unique identification number over
the word NAME on the filter paper card you will use to collect the blood sample from the
respondent. Paste the third bar code label with the same unique identification number on the
blood transmittal sheet (Figure 3.3). This sheet will provide a record of all the blood samples
collected in a cluster for HIV testing.

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 • Do not attempt to peel an incorrect label off of any form or filter paper card. If you make a
mistake with the labels, use the next complete set of labels and paste them over the incorrect
ones.

More detail on this procedure is provided in Section 6, which describes the process of collecting blood for
HIV tests.

3.4 Providing VCT Information

Along with an informational brochure on HIV/AIDS that will be left with the household, a list of
participating VCT centers where vouchers distributed by the AIS can be taken for free HIV Voluntary
Testing and Counseling services will be provided.

All eligible respondents and their spouse/partner, regardless of their participation or non-participation in
the HIV testing, as well as anyone requesting such information should be provided with voucher(s) for
free HIV VCT services, a list of participating VCT centers, and an informational brochure about
HIV/AIDS.

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 Figure 3.3 Blood Transmittal Sheet – Front
┌────┬────┐ ┌────┬────┬────┐
TEAM │░ │░░ │ CLUSTER │░ ░│░ │ │
NUMBER │░ ░│ │ NUMBER │░ ░ │░ │ │
└────┴────┘ └────┴────┴────┘

SIGNATURE SIGNATURE
(CONFIRMING (CONFIRMING NOTES
PERSON TOTAL THAT EACH THAT THE (NOTE ANY
SENDING/ COUNT OF SAMPLE IS NUMBER OF DISCREPANCY
RECEIVING TIME TO FILL BLOOD PRESENT—SEE BLOOD SAMPLES IN NUMBERS
SAMPLES IN THE FORM SAMPLES BACK OF FORM) MATCHES COL. 3) DATE OF SAMPLES)
(1) (2) (3) (4) (5) (6) (7)

INTERVIEWER WHEN ┌────┬────┐

│░░ │ ░ ░│
AND TEAM CLUSTER IS │░░ │ ░ ░│
SUPERVISOR COMPLETED └────┴────┘
WHEN ┌────┬────┐
FIELD SAMPLES ARE │░░ │ ░ ░│
COORDINATOR PICKED UP IN │░░ │ ░ ░│
FIELD └────┴────┘
UPON ARRIVAL
TECHNICAL
AT THE ┌────┬────┐
DIRECTOR OR
CENTRAL │░░ │ ░ ░│
MEDICAL │░░ │ ░ ░│
STATISTICAL
COORDINATOR └────┴────┘
OFFICE
HEAD UPON ARRIVAL ┌────┬────┐
│░░ │ ░ ░│
LABORATORY AT │░░ │ ░ ░│
TECHNICIAN LABORATORY └────┴────┘

INSTRUCTIONS

Interviewer: Upon completion of a cluster, verify that the unique bar code (identification) number on each blood sample (filter paper card)
collected and stored in the large zip-loc bag labeled with that cluster number corresponds to a bar code number pasted to the back of this
transmittal sheet and vice-versa. Note any discrepancies in Column (7). Count and record the total number of blood samples in Column (3). Sign
your name in Column (4) and the date in Column (6). Fold and store this transmittal sheet in the large zip-loc bag.

Field Team Supervisor: After the interviewer has verified the blood samples, you will conduct a second verification. Verify that the unique bar
code (identification) number on each blood sample (filter paper card) collected and stored in the large zip-loc bag labeled with that cluster
number corresponds to a bar code number pasted to the back of this transmittal sheet and vice-versa. Note any discrepancies in Column (7).
Count and verify the total number of blood samples in Column (3). Sign your name in Column (5) and the date in Column (6). Refold and store
this transmittal sheet in the large zip-loc bag.

Field Coordinator: Before returning to the General Statistical Office after visiting a team in the field, you will verify the number of blood
samples collected in each completed cluster that you are carrying back with you. For each completed cluster, count and record the total number of
blood samples stored in the large zip-loc bag labeled with that cluster number in Column (3). Note any discrepancies in Column (7). Sign your
name in Column (5) and the date in Column (6). Refold and store this transmittal sheet in the large zip-loc bag.

General Statistical Office Receiver: For each large zip-loc bag arriving from the field, you will verify the number of blood samples received.
Count and record the total number of blood samples stored in the large zip-loc bag labeled with the cluster number in Column (3). Note any
discrepancies in Column (7). Sign your name in Column (5) and the date in Column (6). Photocopy both sides of this transmittal sheet and file the
photocopies (as instructed) in a designated, locking file cabinet. Refold and store the original transmittal sheet in the large zip-loc bag. Follow
the AIS protocol for sending blood samples to the laboratory for testing.

Laboratory Receiver: Upon receiving blood samples from the General Statistical Office, verify that the unique bar code (identification) number
on each blood sample (filter paper card) collected and stored in the large zip-loc bag labeled with the cluster number corresponds to a bar code
number pasted to the back of this transmittal sheet and vice-versa. Note any discrepancies in Column (7). Count and record the total number of
blood samples in Column (3). Sign your name in Column (4) and the date in Column (6). Follow the AIS protocol for storing and processing
blood samples.

Note: this form will be destroyed under the direction of the Lab Director after all blood samples have been completely processed and a
Final HIV Test Result has been determined for each usable sample.

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 Figure 3.4 Blood Transmittal Sheet – Back

NO. BAR CODE TECH LAB NO. BAR CODE TECH LAB
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:
1 15
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:
2 16
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3 17
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4 18
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:
5 19
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6 20
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7 21
↑
FOLD
.... ........................................ ..... ← HERE → ........................................ ..... .....

↓
8 22

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9 23
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10 24
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11 25
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12 26
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13 27
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14 28
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:

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 Figure 3.5 Sample Sheet of Bar Code Labels

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SECTION 4: PROCEDURES FOR COLLECTING CAPILLARY
BLOOD DROP SAMPLES

4.1 Steps in Obtaining Capillary Blood from the Finger

Capillary blood can be obtained from the palm side of the end of a finger. The following describes the
steps that are involved in obtaining a capillary blood drop sample from the finger.

Step 1: Complete General Preparation

a. If possible, find an indoor site to encourage privacy. If possible, the site should have a table or other
piece of furniture with a flat surface where you can lay out the supplies. A couch, bed or mat should
be readily available should the respondent feel faint and
Table 4.1
need to lie down.
Equipment and Supplies Needed
If you find you must do the test outdoors, find a site in the
For the Finger Prick
full shade and away from rain, dust, and other environmental Retractable lancets for adults
elements that might affect the sample. Latex gloves
Alcohol swabs
b. Take out a plastic bag and spread it over a flat surface Sterile gauze pads
where you will lay out your supplies. Bandaids
c. Confirm the number of eligible respondents from the For the HIV Testing
household questionnaire for whom blood samples will be Filter paper cards
collected. After you have established the number of Bar code labels
respondents to test, take out the appropriate equipment and Drying box
supplies (Table 4.1). You will want to have all materials in
easy reach when you begin collecting blood samples from Other supplies/equipment:
the respondents. Plastic bags for waste

d. When and where possible, wash and dry your hands. Put on gloves before beginning the collection of
the blood sample from the first respondent.

Step 2: Select and Prepare the Puncture Site

a. It is usually easier if you sit on the side of the respondent Puncture sites
opposite to the hand that you will collect blood from. For
example, if you want to collect the specimen from the left
hand, place yourself to the right side of the respondent.

b. Use the third or fourth finger for collecting the blood

(Figure 4.1). Do not use a finger with a scar, a wound or cut,
an infection, swelling, a deformity, or a rash. Also, do not
use a finger on which the respondent is wearing a ring,
because the ring may disrupt the free flow of blood to the tip
of the finger.
Figure 4.1 Fingers for blood collection
c. If the hand is cold, warm the skin over the puncture site by
rubbing it. This will increase blood flow by reducing tissue
fluid and will improve the ease with which a sample can be
obtained.

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d. With an alcohol swab, clean the skin of the finger
thoroughly (Figure 4.2). If the skin is very dirty, use a second
swab. Allow the alcohol to air dry. Do not blow on the area to
dry the alcohol. Blowing may allow bacteria to contaminate
the site.

e. Ensure that the lancet is easily accessible. For adults, you

will use adult lancets which are blue in color and have a 2.4
mm blade.

f. Remove the white blade slot cover by first twisting it 360o and Figure 4.2 Clean the finger
then pulling it out. Do not pull out the needle slot cover
without twisting it first as this may cause the needle not to
pierce the skin. Puncture sites

Step 3: Prick the Finger

a. Make sure that the finger is below the level of the

respondent’s heart to increase the flow of blood to the finger.
Using a rolling movement of your thumb, lightly press the
finger from the top knuckle toward the tip. That action will
stimulate a flow of blood to the sample area.

b. When your thumb reaches the fingertip, maintain a gentle Figure 4.3 Puncture sites on the finger
pressure. Place the lancet perpendicular to the surface of
the end portion of the finger, either centered or slightly off
center. Avoid the very tip of the finger or the sides, because of
the risk of piercing the underlying bone (Figure 4.3).

c. Use the lancet to puncture the skin by placing the blade-slot

surface against the area and pressing the trigger (Figure 4.4).
The tip of the blade ejects through the blade slot, producing a
micro-incision in the skin, and immediately retracts into the
device. After puncturing the skin, turn the finger slightly to
prevent blood from running into the grooves of the
fingerprints. Figure 4.4 Using lancet to prick finger

d. After blood collection is completed, the lancet should be

placed in the plastic bag provided for biohazardous wastes,
along with the other disposable materials used for the blood
sample collection.

Step 4: Collect the blood drops

a. When the blood appears, use a sterile gauze pad to wipe away
the first drop of blood (Figure 4.5). The second, third, and
fourth and fifth drops of blood will be collected on a labeled
filter paper card for HIV testing. Collect a sixth drop if Figure 4.5 Wiping away first blood drop
possible.

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b. If the blood stops flowing before a sufficient amount has been collected, the skin puncture procedure
may be repeated with the respondent’s (parent’s/responsible adult’s) consent on a different finger. Do
not reuse any of the supplies used for the first puncture.

Step 5: Collect biohazardous wastes

Place all biohazardous wastes (e.g., lancets, alcohol swabs, gauze, and gloves) into a plastic bag provided
for field disposal of these items. At the end of the day, follow the procedures described in Section 7 for
the proper disposal of these waste materials.

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SECTION 5: HIV TESTING FOR ADULTS AND YOUTHS

This section of the manual focuses on the various steps involved in collecting the blood spot samples for
HIV laboratory testing.

5.1 Collecting Blood for HIV Testing

The principal tasks involved in HIV testing for adults and youths include:

• Obtain voluntary consent of the respondent;

• Place the bar code labels on a filter paper card, the HIV Testing Page of the questionnaire, and the
Blood Sample Transmittal Sheet;

• Prick the respondent’s finger;

• Collect the second, third, fourth, and fifth blood drops from a finger prick on a labeled filter paper
card;

• Stop the bleeding at the prick site;

• Place the filter paper card from the respondent in the drying box;

• Provide all respondents with an informational brochure on HIV/AIDS, coupons for free VCT
services, and a list of participating VCT centers;

• Collect biohazardous waste.

Procedures for pricking the respondent’s finger were detailed in Section 4. The following describes the
steps that should be followed when collecting blood samples for HIV .

Step 1: Placement of the bar code labels

a. Wearing a pair of latex gloves, carefully remove a new filter paper card from the plastic zip-loc bag in
which you have stored the cards. Make sure to handle the card in such a way that you do not touch the
areas within the preprinted circle. Never handle a card with your bare hands as you may transfer
sweat, dirt or other contaminants on to the card.

b. Put the card with the preprinted circles face-up on the clean absorbent paper sheet that you have
spread out on a flat surface. Discard the card if it drops on the floor or ground or if it becomes dirty in
any other manner.

c. Take the first bar code label from the next full row on the sheet of labels and paste it in Question
705 of the Blood Spot Collection Page for HIV testing.

d. Take the second bar code label from the same row on the sheet of labels and place it at the bottom of
the filter paper card where it says ‘NAME’. Do not cover up any part of the preprinted circles.
Remember not to touch the circles.

e. Take the third bar code label from the same row on the sheet of labels. Paste it on the Blood Sample
Transmittal Sheet for this cluster that will be sent together with the samples to the central statistical
office and then to the laboratory.

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f. DO THIS CAREFULLY. The bar code label is the only means of identifying the blood sample and
for eventually linking the final HIV antibody test results to the interview data. Mistakes will result in
mismatches later on. CHECK THAT ALL THREE LABELS FOR A RESPONDENT HAVE
BEEN PLACED ON THE FILTER PAPER CARD, THE TEST PAGE, AND THE
TRANSMITTAL FORM BEFORE YOU PROCEED TO COLLECT BLOOD DROPS FROM
THE RESPONDENT. If you have more than one eligible respondent from whom all necessary
consents for HIV testing have been obtained, first finish all steps for one respondent before repeating
all steps for the next respondent.

Step 2: Collecting Blood Spots on the Filter Paper Card

a. Follow the instructions in Section 4 for pricking the finger

and wiping away the first blood drop. Be sure to hold the
respondent’s hand at a level lower than his/her heart to
promote blood flow.

b. While maintaining a firm grip on the finger, press gently

on the side of the finger from which you are taking the
blood sample to get a large second drop. Be careful to
avoid ‘milking’ or ‘squeezing’ the finger as this could
affect the test results. Wait until the drop is large enough
to fill one of the preprinted circles on the card (Figure 5.1). Figure 5.1 First drop of blood

c. Move the card underneath the finger. Allow the drop to

fall. If it does not, the filter paper can be touched gently
against the LARGE blood drop (but not the skin) and, in
one step, a sufficient quantity of blood should be
allowed to soak through and completely fill the circle
(Figure 5.2).

The card must not be pressed against the puncture site

on the finger. Make sure that the respondent’s finger
does not touch the card at any point when you are
collecting the blood spots. DO NOT ‘layer’ the sample
in an attempt to fill in the circle. Always use the pre- Figure 5.2 Filling circles on the filter
paper card
printed side of the card to collect the blood spots.

d. To enhance blood flow, gently apply intermittent pressure

to the area surrounding the puncture site to get a third and
fourth drop. Allow sufficient time for a large blood drop to
form before filling a second circle on the filter paper card.
Again, avoid milking or squeezing the finger.

e. There may be times when a drop of blood will not

completely fill the circle. If a circle is not completely
saturated, the next drop of blood may be used to saturate
the circle if the drop is obtained immediately. If there is
any interruption in getting the subsequent drop, you must
begin filling another circle. Layering or application of Figure 5.3 Fully-filled circles without
successive drops of blood to the same printed circle causes layering or caking

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 caking and/or non-uniform concentrations of blood.

f. You must continue to collect drops of blood until you have fully saturated at least four circles on the
filter paper card (Figure 5.3) If the blood flow stops or decreases before you fully saturate the four
circles, you will need to do another finger prick. Whenever this is necessary, you should explain to
the respondent that you were unable to obtain an adequate sample and ask permission to obtain blood
from another finger. Use fresh supplies and a different finger for the second finger prick.

g. Place the card with the blood spots away from other items. Be careful not to drop the card.

Step 3: Stopping bleeding at puncture site

a. After the blood drop collection, wipe any remaining blood from the puncture site with a sterile gauze
pad.

b. After making sure that the blood flow has completely stopped, take an adhesive bandage from its
wrapper and apply it to the puncture site.

Step 4: Placing the filter paper card in the drying box

a. The drying box should always be placed vertically on a flat surface before opening. Open the drying
box. Carefully pick up the filter paper card with the blood spots and place it in a horizontal position in
one of the slots in the drying rack in the box. Avoid touching or smearing the blood spots on other
cards in the box when you are storing a new card. Never put more than one filter paper card in a
single slot in the drying rack. As described in Section 2, blood spots must dry overnight at ambient
temperature (15˚C to 22˚C). Never set any materials on top of the open box as they might
contaminate the filter paper cards that are stored there.

b. If the collection process in a household is interrupted for any reason, you should close the box to
prevent any possible contamination.

c. After all blood spot samples for the household have been collected, carefully close the box and return
it to a vertical position. The drying box should be kept in a vertical position at all times so that the
filter paper cards remain in a horizontal position in their slots inside the box.

Step 5: Collecting biohazardous waste

Place all biohazardous waste (e.g., lancets, alcohol swabs, gauze, and gloves) into a plastic bag provided
for field disposal of these items. At the end of the day, follow the procedures described in Section 7 for
the proper disposal of these waste materials.

5.2 Providing Referrals for HIV VCT Material

All eligible respondents and their spouse/partner, regardless of their participation or non-participation in
the HIV testing, as well as anyone requesting such information should be provided with voucher(s) for
free HIV VCT services, a list of participating VCT centers, and an informational brochure about
HIV/AIDS.

5.3 Storing and Transferring the Dried Blood Spots

The dried blood spot (DBS) samples must be carefully maintained until they are picked up and taken to
the laboratory. They should never be exposed to sunlight during storage, and it is important to regularly

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monitor the levels of humidity in the storage box(es). The following describes the steps that should be
followed in storing and transferring the DBS samples.

Step 1: Storing the DBS Samples

Each morning, before you go to the field, you must remove the filter paper cards with the blood spots that
you collected on the previous day from the drying box and prepare them for storage. The following
describes the procedures in this step.

a. Put on a pair of latex gloves and carefully open the

drying box. Check that the blood spots on each
filter paper are completely dried (chocolate
brown).

b. Separately remove each filter paper card on which

the spots have dried from the drying box. Be
careful not to touch the blood spots.

c. Gently fold a piece of glassine paper over the

blood spots and put the filter paper card into a
small (low gas-permeable) zip-loc bag (Figure
Figure 5.4 Supplies for storing the filter paper
5.4). Put one desiccant packet and a humidity cards
indicator card behind the filter paper card, facing
out so that the humidity level is visible. It is important that the desiccant packet and the humidity
indicator card do not touch the blood spots. Close the zipper, gently pushing out any excess air in the
bag as you are zipping it, being careful not to press on the blood spots. DBS samples should not be
allowed to come into contact with other DBS samples during handling, shipment or storage.

d. Continue to package each of the filter paper cards from the previous day which have dried overnight,
putting each one into a bag with a desiccant packet and a humidity indicator card. When you have
packaged all of the filter paper cards, put them into the large zip-loc bag that has been labeled for the
cluster in which the samples were collected. Note that the Cluster Sample (large zip-loc) Bag itself
should also contain a few loose desiccant packets and a humidity indicator card.

e. Every couple of days, before adding newly packaged DBS samples to an existing Cluster Sample
Bag, check the humidity indicator cards for the individually packaged DBS samples that you have
previously placed in the Cluster bag. The buildup of humidity can damage the quality of the
sample.

A bottom circle that is pink (30%

humidity) indicates a warning of
increasing humidity. If the middle circle
(40%) or top circle (50%) is pink, gently
open the small zip-loc bag, remove the
desiccant packet and replace it with a
fresh desiccant packet. If any of the
circles on the humidity indicator card
have merged together so that they are
not completely separated, remove the
indicator card and replace it with a fresh
indicator card. Close the zipper, gently

AIDS Indicator Survey Figure 5.5 Packaged DBS filter paper cards
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 pushing out any excess air in the bag as you are zipping it (Figure 5.5).

Before closing the zipper on the Cluster Sample Bag, check the condition of the packets and humidity
indicator card that are kept loose inside the bag and replace as needed.

If you have additional Cluster Sample Bags for completed clusters that have not yet been collected by
a Field Supervisor, examine all of the samples in those bags in the same manner every couple of days,
as long as they are with your team in the field.

Step 2: Transferring the DBS Samples

The purpose of the Blood Sample Transmittal Form is to account for the samples at each step of the way.
Fold the Blood Sample Transmittal Form along the dotted lines (so that the bar-code labels are not
folded), and keep it in the Sample Cluster Bag along with the DBS samples for that cluster.

When you have completed the cluster, remove the packaged DBS samples from the Cluster Sample Bag
(do not open the small zip-loc bags). One by one, check the bar codes on the labels on the filter paper
cards against the bar codes affixed to the back side of the Blood Sample Transmittal Sheet. For each DBS
sample, put a check mark in the column labeled TECHNICIAN for each corresponding bar code found on
the transmittal sheet. Count the number of DBS samples and record in the boxes provided in Column (3)
on the front side of the Transmittal Form in the row labeled TEAM LEADER. If there are any
discrepancies, you must attempt to account for them. Use Column (7) to explain. Sign your name in
Column (4) and the date in Column (6).

The team’s field supervisor will follow behind you, verify the samples, and sign his name in the same
row.

Periodically, a Field Coordinator or other staff member the central office will visit the teams to collect the
completed questionnaires and the DBS samples for the completed clusters. When he/she collects the DBS
samples, he/she will recount the DBS samples for each of the completed clusters and sign the Blood
Sample Transmittal Form. The samples and transmittal sheet will be transported to the central survey
office for logging before being transferred to the laboratory for processing.

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SECTION 6: PRECAUTIONS TO OBSERVE WHEN
COLLECTING BLOOD SAMPLES

This section of the manual reviews some of the major precautions that you must observe in collecting
samples, both to protect themselves yourself and the subjects from which they you are collecting samples
from injury or infection and to prevent contamination of the samples.

6.1 General Precautions when Collecting Blood

This section describes the universal (general) precautions to be followed during blood collection for HIV
testing. The person responsible for collecting blood for HIV testing must take precautions to prevent
parenteral, skin, and mucous-membrane exposures to blood-borne infections, such as hepatitis B, or
human immunodeficiency virus (HIV). Under general precautions the following rules should be followed
to ensure protection from acquiring blood-borne infections1.

• Wear gloves. Gloves help to prevent skin and mucous-membrane exposure to blood. Gloves
should be worn during blood collection until the specimen(s) from a subject is collected and all
waste materials produced during the collection are disposed of. At that point, the gloves used with
the subject should be treated as biohazardous waste. A new pair of gloves should be used with
each subject. Gloves must never be reused!

• Avoid penetrating injuries. Although gloves can prevent blood contamination of intact and non-
intact skin surfaces, they cannot prevent penetrating injuries caused by the instruments used for
finger or heel pricks. Self-retractable lancet devices reduce the risk of penetrating injuries.

Lancets should not be used for purposes other than a single finger prick to collect blood for HIV
testing. The lancets should not be broken or destroyed for curiosity or other purposes.
Immediately after the testing is completed, the devices should to be placed in a puncture-resistant
disposal biohazard bag.

If an accident occurs, any skin surfaces or mucous membranes that become contaminated with
blood should be immediately and thoroughly washed.

• Never eat or drink during the testing. Since eating, drinking, and applying cosmetics may
distract you from performing the procedure, they are not permitted during HIV blood collection.

• Properly dispose of all biohazardous materials. All materials coming in contact with blood
must be placed in a biohazardous waste container after use and disposed of according to the
survey’s organization’s policy on infectious waste disposal (see Section 7). Take precaution when
storing and transporting the waste container during the fieldwork.

6.2 Rules for Collecting Specimens for HIV Testing2

There are a number of specific rules or precautions that the you must observe when collecting the samples
for HIV testing. These include:

1
For the universal precautions regarding blood-borne pathogens, see the U.S. Centers for Disease Control and
Prevention guidelines and the U.S. Occupational Safety and Health Administration (OSHA) standards for protection
from exposure to blood-borne pathogen.
2
Adapted from NCCLS 1997

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 • Never “milk” the finger. Excessive massaging or squeezing of the finger or foot will cause
tissue juice to mix with and dilute the blood. This may result in erroneous test results. Instead, the
tester should employ only mild pressure by using the thumb and the second and third fingers to
make a “pad” at the puncture site. This will make the connective tissue underlying the skin more
porous and allow the capillary blood to flow easily after the incision.

• Never mix alcohol with the blood. Alcohol, which is used to clean the puncture site, can mix
with the blood and cause errors in the testing results. To avoid this problem, the finger must be
wiped dry completely before being punctured.

• Avoid obstructing blood flow. It is important to hold the finger properly to allow for the
accumulation of blood in the puncture-site area. Holding the finger too tightly can obstruct the
blood flow to the finger.

• Avoid shallow punctures. A deep puncture should be made for better blood flow and to have a
representative concentration of red blood cells.

• Do not layer. Do not layer multiple drops of blood on the target spot (pre-printed circle on the
filter paper card). If blood flow diminishes to incompletely fill circles, request consent from the
subject (parent/guardian) to repeat the sample collection again using another finger.

• Never touch circles on the filter card. Avoid touching the area within the preprinted circles on
the filter paper card before collection. Never touch the blood spots after collection.

• Protect the filter card from contamination. Do not allow water or other contaminants to come
into contact with the specimen card before or after use.

• Do not replace the specimens in the transport container until thoroughly dry (chocolate
brown). Insufficient drying adversely affects test results.

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SECTION 7: BIOHAZARDOUS WASTE DISPOSAL

Any material coming in contact with blood or serum (lancets, alcohol swabs, gauze, and gloves) is
considered to be biohazardous, i.e., hazardous to other humans. Safe disposal of such material is very
important to prevent the transmission and spread of various blood-borne diseases, such as hepatitis B and
HIV, among survey personnel and within the study community. Biohazardous waste has to be collected in
a special container during the blood collection and testing, securely stored and transported, and safely
disposed at the end of each day of fieldwork.

If possible, commercially available biohazardous waste disposal containers should be used for waste
disposal. These types of containers are red and have a special logo warning about biohazardous content.
They can be securely closed for safe storage and transportation during the fieldwork. Whenever possible,
these containers should be taken to health facilities, which employ standard procedures for biohazardous
waste disposal.

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