TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA

SCREENING OF HbsAg, Anti-HIV AND

TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

APPROVED BY:

…………………………………...…………
ASSOC PROF DR ROSLINE HASSAN
HEAD OF HAEMATOLOGY DEPARTMENT

CONTROLLED COPY NO : 4

REGISTERED HOLDER

TRANSFUSION MEDICINE LABORATORY

RECORD OF REVIEW/AMMENDMENT

DATE VERSION NO. DETAIL OF AMMENDMENT BY

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 TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA
SCREENING OF HbsAg, Anti-HIV AND
TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

PREPARED BY : ROHANA YUSOF

DESIGNATION : TECHNICAL OFFICER / MEDICAL LAB TECHNOLOGIST U32

CHECKED BY : DR. MARINI RAMLI

DESIGNATION : HAEMATOLOGIST

AUTHORISED BY : ASSOC PROF DR ROSLINE HASSAN

DESIGNATION : LAB DIRECTOR / HEAD OF DEPARTMENT

1. OBJECTIVE

To screen all donated blood in order to ensure the safety of donor blood from Hepatitis
B, HIV and Hepatitis C infection.

2. METHOD

Microparticle Enzyme Immunoassay (MEIA) technology for the qualitative detection of

Hepatitis B Surface Antigen (HBsAg), antibody to hepatitis C virus (anti-HCV) and antibodies to
human immunodeficiency virus type 1 and/or type 2 (HIV 1/HIV2) in human serum or plasma.

3. PRINCIPLE

MEIAs are a variation of the enzyme immunoassay (EIA) principle. Solid phase EIAs
use antigens and/or antibodies coated on a surface to bind complementary analytes. The
bound analyte is detected by a series of antigen-antibody reactions. In the AxSYM final
reaction, an antibody coupled to an enzyme acts upon a substrate to produce a fluorescent
end-product. The fluorescence produced by the enzyme reaction is measured by MEIA optical
assembly. The presence or absence of HBsAg, anti-HIV and anti-HCV is determined by
comparing the rate of formation of fluorescent product to the cutoff rate which is calculated from
a previous AxSYM Index Calibration. If the rate of formation of fluorescent product in the
sample is greater than or equal to the cutoff rate, the sample is considered reactive.

(Refer AxSYM System Operations Manual)

4. REQUIREMENTS

4.1 Equipment:
AxSYM Analyser
Centrifuge

4.2 Disposable Pipettes

Sample cups
Plain test tubes
Matrix Cells
Reaction Vessels

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 TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA
SCREENING OF HbsAg, Anti-HIV AND
TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

4.3 Reagents
Reagent packs – AxSYM HBsAg (V2), AxSYM HIV 1/2 gO, AxSYM HCV version
3.0,
Reagent controls for AxSYM HBsAg (V2), AxSYM HIV 1/2 gO, AxSYM HCV version
3.0,
Index calibrator
Probe Cleaning Solution,
Solution 1 (MUP)
Solution 3 (Matrix Cell Wash)
Solution 4 (Line Diluent)

4.4 Donor serum or plasma

5. PROCEDURE

NO ACTIVITY RESPONSIBILITY
5.1 Preparing samples MLT
5.1.1 Receive donor samples with ‘Borang Pendaftaran
Penderma Darah’ (BPPD) and ‘Borang Penyerahan
Darah Penderma Untuk Ujian Serologi’ (BPDP)

5.1.2 Check identity of samples with information on BPPD. If

any discrepancy exists and condition of blood samples
unsuitable (eg: total numbers of samples not tally with
BPPD, hemolysed, inadequate, no barcode, etc.), resolve
accordingly by obtaining correct sample from blood bag,
inform officer in-charge and record in BKUS (Refer
Appendix 1)

5.1.3 Allow the samples to stand for at least two hours before
processing if using plain tube without gel separator.

5.2 Processing sample MLT

5.2.1 Centrifuge the donor samples at minimum 1520 x g for 15
minutes to separate the serum

5.2.2 Arrange the plain tube into suitable sample segment

according to blood bag number.

5.2.4 Ensure all samples are free from air bubbles and foaming.

5.3 Initialize the AxSYM analyzer MLT

5.3.1 Check the inventory and perform machine maintenance
each run (Refer Appendix 2).

5.3.2 Perform reagent calibration on all new reagent lot number

or as required. (Do not simultaneously calibrate more
than one reagent lot).

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 TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA
SCREENING OF HbsAg, Anti-HIV AND
TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

5.3.3 Run control daily or as required.

i) Perform internal quality control as follow:
- AxSYM HBsAg: Negative Control and Positive
Control
- AxSYM HIV 1/2 gO: Negative Control, HIV-1 Positive
Control and HIV-2 Positive Control
- AxSYM HCV: Negative Control and Positive Control.

ii) Verify that the AxSYM control values are within

acceptable ranges (Refer Appendix 3). If one of the
AxSYM Control values is out of its specified range,
troubleshoot and perform recalibration.

5.3.4 Perform HBsAg, anti-HIV and anti-HCV assays

(Refer AxSYM System Operation Manual)

5.4 Results MLT

5.4.1 Ensure that results produced are tallied with ordered list
of samples according to arrangement.

5.4.2 Print the control and test results. Make sure all results are
released and saved.

5.4.3 Transfer the results interpretation into ‘Borang Keputusan

Ujian Saringan’ (BKUS). Recheck all the results to make
sure no discrepancy.

5.5 Initially Reactive (IR) repeat test MLT

5.5.1 Select Initial Reactive test results.

5.5.2 Label blood bag barcode number and reactive assay on

plain test tube using permanent felt tip marker.

5.5.3 Get the IR sample from tested plain tube, examine for clot
and transfer sample to the labeled plain test tube.

5.5.4 Isolate Initial Reactive Blood Bag (IRBB). Check IRBB

barcode number to tally with the IR barcode number in the
printed result list.

5.5.5 Transfer sample from the blood bag into another labeled
plain test tube.

5.5.6 Centrifuge both the plain test tubes at minimum 10,000 x

g for 10 minutes.

5.5.7 Transfer serum into new labeled sample cups. Repeat the
particular assay on the samples

5.5.8 Check for any results discrepancy. If there is any

discrepancy, resolve accordingly. (Refer Appendix 4)
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 TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA
SCREENING OF HbsAg, Anti-HIV AND
TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

5.5.9 Verify the test results by signing initial on every page.

5.5.10 Collect and store repeatedly reactive sample from

segment of reactive blood bag in labeled plain test tube
and keep serum sample in 2-8ºC for further confirmatory
test as follow:
i) antiHCV: Inno-LIA HCV
ii) antiHIV: Particle Agglutination test and Western Blot
5.5.11 Send the sample for confirmatory test to Microbiology
Laboratory.

5.6 Reporting results:

5.6.1 Print the repeated test results and record result into MLT
‘Borang Keputusan Ujian Saringan Kedua’ (BKUS2)
(Refer Appendix 5). Verify the test results.

5.6.2 Check and sign all results in BKUS and BKUS2 by signing MLT U38/
initial on every page. U36/U32

5.6.3 Verify and sign all checked results. SO

5.6.4 Enter all results interpretation in TMSISS system. Print MLT

and verify TMSISS printed results are tallied with AxSYM
printed results. Sign the TMSISS printed reported results.
File in ‘Rekod Penderma Darah dan Keputusan Ujian
Saringan’.

Note: For releasing of emergency results in case of shortage of

blood, senior technologist on-call is required to check the
results before the blood are released. Scientific Officer on-call
will be informed for approval of releasing the results.

5.7 Discard Blood Bag and Blood Components MLT

5.7.1 Search for all reactive blood bag and its blood
components.

5.7.2 Tally both barcode number between the blood bag and all
components prepared with BKUS2.

5.7.3 Record in file ‘Rekod Pelupusan Darah/Komponen

Tercemar’.

5.7.4 Keep the tainted blood in ‘Komponen Darah Tercemar’

fridge. Autoclave and dispose in yellow biohazard bag for
weekly disposal (Refer HUSM/HEMA-UPT/QP-02, Appendix
3).
5.8 Discard specimen and samples MLT
5.8.1 Discard all tested samples in plain tube after one week.
Record in ‘Buku Rekod Pelupusan Bahan Biological oleh
Radicare’.
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 TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA
SCREENING OF HbsAg, Anti-HIV AND
TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

5.8.2 Reactive samples in sample cup/plain test tube (pilot and

segment) will be kept stored frozen (-10ºC or colder) in
specified box according to the particular assay for one
year.

6. LIMITATION

6.1 Specimens containing red blood cells may give inconsistent results, and therefore
must be centrifuged prior to the testing.

6.2 Centrifuged specimens with a lipid layer on top of the liquid must be transferred to
a sample cup or secondary tube. Care must be taken to transfer only the clarified
specimens and not the lipemic material.

6.3 Frozen specimens and those containing particulate matter must be centrifuged
prior to running the assay.

6.4 Do not use Solution 1 (MUP) beyond the expiration date or maximum of fourteen
days on-board the AxSYM System. Otherwise, it must be stored at 2-8ºC. When
loading new Solution 1 (MUP), it is important to tighten the instrument cap for MUP
to minimize exposure to air. Prolonged exposure to air may compromise
performance.

6.5 Do not use kits beyond the expiration date or a maximum of 112 cumulative hours
on-board the AxSYM System.

6.6 Do not mix reagents from different reagent packs and do not mix reagent packs
and index calibrators from different lots.

6.7 All samples (donor samples, controls and calibrators) should be tested within 3
hours of being placed on-board the AxSYM System.

7. REFERENCE RANGE

7.1 Specimens with a S/CO value less than 1.00 are considered nonreactive
7.2 Specimens with a S/CO value equal to or greater than 1.00 are considered
initially reactive
7.3 Instrument grayzone reading for S/CO* is set as follow:
- HBsAg and anti-HIV assay: 0.9-0.99
- Anti-HCV assay: 0.8-0.99
*(S/CO= Sample Rate/Cutoff Rate)

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 TRANSFUSION MEDICINE LABORATORY, HOSPITAL UNIVERSITI SAINS MALAYSIA
SCREENING OF HbsAg, Anti-HIV AND
TITLE : VERSION NO. 1
Anti-HCV ON BLOOD DONORS

PROCEDURE NO. HUSM/HEMA-UPT/STM-S1 VERSION DATE. 01.11.2009

8. RESULT

8.1 The AxSYM System calculates the cutoff rate (CO) from the mean rate of Index
Calibrator replicates and stores the result. The AxSYM assay calculates a result based
on the ratio of sample rate (S) to the cutoff rate for each sample and control
(S/CO= Sample Rate/Cutoff Rate)

POS interpretation: Reactive

NEG interpretation: Negative / Nonreactive
GZ interpretation: Grayzone

8.2 Back up of Test Results

All test results are to be kept as back up in the diskettes and treated as confidential. The
diskettes are to be kept in the transfusion microbiology scientific officer room

9. REFERENCES

9.1 AxSYM HBsAg (V2) package insert manual

9.2 AxYSM HIV 1/2 gO package insert manual
9.3 AxSYM HCV version 3 package insert manual

END OF DOCUMENT

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