SOP For Cryptococcus Test

Ambo General Hospital Laboratory
Standard Operating Procedure

For
Cryptococcus Lateral flow assay for the detection for
Cryptococcal antigen
Compiler: Signature:
Approver: Signature:
Effective Date:July 01, 2017 Version No. 02
Document No: DGHL/HEM/SOP5.5-12 Copy No. _____
Revision and amendment

Ambo General Hospital Laboratory Document No: AGHL/ SERO/SOPs 5.5-xx
Version No:01
SOP for CrAg Lateral flow assay Page 1 of 7
for the detection for Cryptococcal Effective Date: January2, 2024
antigen
A. Annual Review of Document
Revision Reviewed by: Approved by:

Review Date
No. Name Signature Name Signature
1 Jan2,2024 Bedilu Bogale Geleta
B. Version Change History/Description

Version. Effective Description of Version Change Name & Name &
No. Date Signature of Signature of
Reviewer approval
C. Amendment
Rev. Page Description of Amendment Amendment Effective Name &
No. No Date Date Signature of
approval
Authority of issue: DGHL Authorized date: January1,2024

This is a controlled document for internal use only
Version No:01
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Purpose The CrAg Lateral Flow Assay is an immunochromatographic test system for the
qualitative or semi-quantitative detection of the capsular polysaccharide antigens of
Cryptococcus species complex (Cryptococcus neoformans and Cryptococcusgattii)
in serum and cerebrospinal fluid (CSF).
Abbreviations CrAg--------------Cryptococcus antigen

CSF---------------Cerebrospinal fluid
LF ---------------Lateral Flow
Materials Supply and Material

A. LF Specimen Diluent (2.5 ml, GLF025): Glycinebuffered saline containing
blocking agents and a preservative
B. CrAg LF Test Strips (50 strips in desiccant vial, LFCR50)
C. CrAg Positive Control (1 ml, CB1020): Glycine- buffered saline spiked
with cryptococcal antigen
D. LF Titration Diluent (6 ml, EI0010): Glycine-buffered saline containing a
preservative
E. Package insert
F. Pipettor (40-µL and 80-µL)
G. Timer
H. Disposable micro-centrifuge tubes, test tubes, or a micro-titer plate
I. Glove
J. Syringe
K. Glove
Reagents Storage
All reagents included in this kit should be stored at room temperature (22-25°C) until
the expiration dates listed on theReagent labels.
Special Safety Refer to __________ Referral Hospital Laboratory health and safety guideline.
Precautions
SPECIMEN For optimal results, sterile non-Hemolyzed serum or CSF should be used. If a delay
handling and is encountered in specimenprocessing, storage at 2-8°C for up to 72 hours is
storage permissible.Specimens may be stored for longer periods at <-20°C,provided they
are not repeatedly thawed and refrozen. Specimens in transit should be maintained
at 2-8°C or <20°C.

Procedure Step Action
Ambo General
1 Hospital
Add 1 dropLaboratory
of LF SpecimenDocument No: AGHL/
Diluent (GLF025) SERO/SOPs
to an 5.5-xx
appropriate container
(micro-centrifuge tube, micro titerplate, test tube, etc.).
Version No:01
antigen
2 Add 40 µL of specimen to the container.
3 Submerge the white end of a Cryptococcal Antigen Lateral Flow Test Strip
(LFCR50) into the specimen.
4 Wait 10 minutes.
5 Read and record the results (See READING THE TEST).

Version No:01
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Principle The CrAg Lateral Flow Assay is a dipstick sand wich immunochromatographic assay.
Specimens and specimendiluent are added into an appropriate reservoir, such as a
test tube, and the lateral flow device is placed into the reservoir. The test uses specimen
wicking to capture goldconjugated, anti-CrAg monoclonal antibodies and
goldconjugated control antibodies deposited on the test membrane. If CrAg is present
in the specimen, then it binds to the gold-conjugated, anti-CrAg antibodies. The
goldlabeled antibody-antigen complex continues to wick up the membrane where it will
interact with the test line, which has immobilized anti-CrAg monoclonal antibodies.
The goldlabeled antibody-antigen complex forms a sandwich at the test line causing a
visible line to form. With proper flow and reagent reactivity, the wicking of any
specimen, positive or negative, will cause the gold-conjugated control antibody to move
to the control line. Immobilized antibodies at the control line will bind to the gold-
conjugated control antibody and form a visible control line. Positive test results creat
two lines (test and control). Negative test results form only one line (control). If a
control line fails to develop then the test is not valid.
Clinical For screening of Cryptococcal infection from the patient specimen.

Utility
1. Doering, T. L. 2009. How sweet it is! Cell wall biogenesis and polysaccharide
Reference capsule formation in Cryptococcus neoformans. Annu.Rev.Microbiol. 63:223-247.
2. 2. Goodman, J. S., L. Kaufman, and M. G. Koenig. 1971. Diagnosis of
cryptococcal meningitis. Value of immunologic detection of cryptococcal antigen.
N.Engl.J.Med. 285:434-436.
3. 3. Kozel, T. R. 1995. Virulence factors of Cryptococcus neoformans. Trends
Microbiol. 3:295-299.
4. 4. Lin, X. and J. Heitman. 2006. The biology of the Cryptococcus neoformans
species complex. Annu.Rev.Microbiol. 60:69-105.
5. 5. Lindsley, M. D., D. W. Warnock, and C. J. Morrison. 2006. Serological and
Molecular Diagnosis of Fungal Infections, p. 569-605. In B. Detrick, R. G.
Hamilton, and J. D. Folds (ed.), Manual of Molecular and Clinical Laboratory
Immunology. ASM Press, Washington, D.C.
6. 6. Park, B. J., K. A. Wannemuehler, B. J. Marston, N. Govender, P. G. Pappas, and
T. M. Chiller. 2009. Estimation of the current global burden of cryptococcal
meningitis among persons living with HIV/AIDS. AIDS 23:525-530.
7. 7. Zhou, Q. and W. J. Murphy. 2006. Immune response and immunotherapy to
Cryptococcus infections. Immunol.Res. 35:191-208.

Version No:01
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Ambo General Hospital Laboratory

Standard Operating Procedure

Effective Date:July 01, 2017 Version No. 02

Document No: DGHL/HEM/SOP5.5-12 Copy No. _____

Revision and amendment

A. Annual Review of Document

Revision Reviewed by: Approved by:

B. Version Change History/Description

Authority of issue: DGHL Authorized date: January1,2024

Abbreviations CrAg--------------Cryptococcus antigen

Materials Supply and Material

Authority of issue: DGHL Authorized date: January1,2024

2 Add 40 µL of specimen to the container.

5 Read and record the results (See READING THE TEST).

Clinical For screening of Cryptococcal infection from the patient specimen.

Authority of issue: DGHL Authorized date: January1,2024

Authority of issue: DGHL Authorized date: January1,2024

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