Product specification
HBsAg latex reagents
1.Product codes and kit sizes
HB/01050 Test kit
HB/012100 Test kit
2.Application and test principle
The HBsAg latex test is a rapid (5 minute) slide agglutination procedure for the detection of Hepatitis B Surface Antigen (HBsAg) in serum. The assay is performed by testing a suspension of latex particles coated with mouse monoclonal antibodies to HBsAg against unknown serum samples. Samples showing distinct agglutination after 5 minutes, indicate a level of HbsAg between 30 and 3,200,000 U/ml based on the Paul Ehrlich institute standard.
3. Manufacturing procedure
Latex particles are coated with mouse monoclonal antibodies to HBsAg, the final product is suspended in a buffered solution to preserve activity. The product is then passed through quality control and once approved for use is dispensed into finished kits as required. All procedures are carried out in strict conformance to ISO9001:2000 guidelines.
4.Test procedure
For full details refer to the Methodology Sheet .
5.Composition of reagents:
Latex Reagent: A suspension of polystyrene latex particles coated with mouse monoclonal antibodies to HBsAg in a buffered solution. Contains less than 0.1% of sodium azide as a preservative.
Positive Control: Heat inactivated human sera with a titre greater than 640 U/ml. Sterile, 0.22 um filtered. Contains less than 0.1% of sodium azide as a preservative.
Negative Control: Human negative sera. Sterile, 0.22 um filtered. Contains less than 0.1% of sodium azide as a preservative.
All sera are tested and found negative for HIV, and HCV by FDA approved methods.
6.Specimen collection and preparation
For full details refer to the Methodology Sheet in appendix (17.3).
7Calculations required
The level of HBsAg in the sample is calculated as the highest dilution to show agglutination when the recommended technique is used, multiplied by 30 (The lower limit of sensitivity of the latex is 30U/ml).
Example: Positive agglutination at titre of 1/40 => 40 x 30 => 1200 U/ml.
8.Interpretation of results
For full details refer to the Methodology Sheet in appendix (17.3).
9. Reference or standard values
The presence of agglutination indicates a level of HBsAg equal to or greater than 30U/ml. The absence of agglutination indicates a level of HBsAg less than 30U/ml.
The sensitivity of the reagent is referred to the Paul Ehrlich standard.
10. Limitations of procedure, interfering substances and precautions
Positive reactions can occur in conditions other than Hepatitis such as mononucleosis, Rheumatoid Arthritis, syphilis and various other infections in elderly patients. The incidence of false positive results in apparently healthy individuals is less than 5%.
The upper detection limit was assessed using three different lots of HBsAg latex reagent using a partially purified preparation derived from a pool of donors. This showed a sample containing 80,000 U/ml gave a positive reaction. Since a dilution of 1:40 of the specimen is tested in conjuction with the specimen itslf, the upper detection limit was calculated as 3,200,000 U/ml.
Bacterial contamination of controls and specimens as well as the freezing and thawing of the antigen may lead to false positive results.
The HBsAg latex reagent must not be used beyond its expiry date as prolonged storage can affect the sensitivity of the suspension.
The sensitivity of the test may be reduced at low temperatures, all tests should be carried out between 15 and 25 oC
A delay in reading the results may result in the over-estimation of the antigen, if present.
Do not dilute any of the products prior to use.
Do not touch the circles of test cards as this may prevent the spreading of the sample.
11. Quality control
All internal quality control is undertaken in a purpose built QC facility. All procedures are carried out in strict conformance to ISO 9001:2000 guidelines.
11.1 Physical - chemical parameters
Appearance: A suspension free of macroscopic or flaky particles.
Colour:White coloured latex.
Settling:No visual evidence of excessive settling of latex particles within 24 hours of suspension.
Bacteriological laod: Less than 2 CFU’s per ml at +37C for 5 days on blood agar plates.
pH:8.20 +/- 0.2
11.2 Sensitivity against in-house standards
The in-house standard is calibrated against a known Paul Ehrlich International Standard.
STANDARD: LOT 15 TITER: 160U/ml
`160804020100 U/ml
RESULT 3+3++---
11.3 Reproducibility
All reagents are tested against three previous batches of test reagents by two different operators, no significant differences were observed.
11.4 Specificity – Immunological reaction
Positive sera: Must give a clear positive reaction (visible agglutination) within 5 minutes with a panel of 50 samples of sera with differing titres.
Negative sera: Must give a clear negative reaction (no visible agglutination) after 5 minutes with a panel of 50 serum samples from normal individuals.
12.Stability standards
12.1 Real time stability
Three different batches were tested every 3 months for the duration of the shelf life – no significant decrease in reaction was observed in any batch.
12.2 Accelerated stability studies/shipping trials
Three different batches were maintained at 45 oC and tested over a period of 5 days - no significant decrease in reaction was observed in any of the batches tested.
13.Storage instructions
All reagents should be stored upright at 2-8 C, under these conditions the products will maintain their activity until the expiration date of the test kit. After use all vials should be tightly closed and stored as above. The components of the test kit are stable until the expiry date which is printed on the vial label. False positive results may occur if any of the reagents have been contaminated.
UNDER NO CIRCUMSTANCES SHOULD THE LATEX REAGENTS BE FROZEN.
14.Disposal of used products
Any materials derived from human origin should be treated as potentially hazardous and handled and disposed of accordingly. No evidence has been found to suggest that any of the kit components or derivatives are toxicological. Care should be taken when products are disposed of, as azides can form explosive salts with copper or lead in plumbing systems. On disposal of products, flush with large quantities of water.
15.Precautions:
For In Vitro diagnostic use only.
All sera are tested and found negative for HIV, and HCV by FDA approved methods.
16.References
C. Mcdonald and J. Barbara, Medical Laboratory Sciences (1988) 45, 277-288
Finny GJ., John R., Babu PG., Tans. R. Soc. Trop. Med. Hyg. (1996), 5: 533-534
Seidl S., Trautmann L., Rev. Fr. Transfus. Immunohematol (1981), 24 (3):319-335