CRP LATEX TEST is a qualitative screening and semi-quantitative determination of C-Reactive Protein antibodies (CRP) in serum.C-RP usually appears in the sera of patients in the acute stages of a number of inflammatory conditions such as most viral and some bacteria infections; rheumatoid arthritis, acute rheumatic fever with or without carditis and most other collagen diseases and in a number of other conditions characterized by inflammation.
CRP is considered to be a sensitive indicator of acute inflammation. Changes in the serum level of CRP with time from the same patient can be used to monitor recovery.The use of the CRP latex test to measure the effectiveness of therapy is of great clinical significance in cases such as acute rheumatic fever.
Principle of CRP LATEX TEST
CRP Latex test is based on latex-agglutination method.That is the immunological reaction between CRP as an antigen and the corresponding antibody coated on the surface of biologically inert latex particles.
SPECIMEN COLLECTION AND PREPARATION
This test should be performed using fresh serum specimens only.
In case of delay,the specimens should be refrigerated (or frozen where applicable) because bacterial contamination may cause protein denaturation resulting to false positive agglutination.
Reagent required for CRP Test (CRP Test Kit)
|CRP Latex Reagent||This contains polystyrene latex particles coated with anti-human CRP in a stabilized buffer with less than 0.1% sodium azide as preservative|
|CRP Positive Control||Made up of human Serum that contains more than 6 mg/L CRP and less than 0.1% sodium azide as preservative.|
|CRP Negative Control||Made up of human serum that has been diluted and stabilized with buffer and contain less than 0.1% sodium azide as preservative.|
|Glycine-saline Buffer: (20X) Concentrate||To be diluted 1:20 with distilled water|
Materials CRP Test
- Disposable pipettes
- test slides
- Physiological saline
- test tubes
- Fresh Serum sample
How to carryout CRP Qualitative Test
Start by bringing all test reagents and serum specimens to room temperature.
- Now gently shake or agitate the CRP latex vial or CRP Kit to disperse and suspend latex particles for at least 10 seconds.
- Positive and negative controls should be tested with each series of test.
- Pipette one drop (50µl) of test serum onto Test,Positive and Negative control circle on the slide
- Pipette one drop of CRP Latex reagent,Positive and negative control to each circle that contains specimens on the slide respectively.
- Spread the resulting mixture by using the paddle end of the pipette or and applicator stick.
- Do not use the same paddle end or applicator stick to mix each test serum or control as this will cause cross-contamination.
- Now gently tilt and rotate slide by hand for two (2) minutes.
- Observe for macroscopic clumping using the indirect oblique light source.
- Compare the reaction of the test serum to the CRP positive and negative control sera.
Observation and Result for Qualitative test
|Negative Result||Smooth milky suspension|
CRP Quantitative Test
Sera that are positive in the screening test (Qualitative test) should be retested in the titration test to provide verification for borderline interpretations. The greatest dilution of test sample showing agglutination is considered the endpoint.
Since negative results may be caused due to excess antigen, the test should be repeated using a diluted serum sample if prozone effect is suspected.
CRP Quantitative Test Procedure
- For each test serum to be titrated, set up a least 6 test tubes and label them as 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, etc.
- To each tube add 0.2 ml of Diluted Glycine-Saline Buffer.
- To Tube No. 1 (1:2) add 0.2 ml of undiluted test serum.
- Serially make two-fold dilutions by mixing contents of Tube No. 1 with pipette and transferring 0.2 ml to Tube No. 2.
- Repeat serial transfers for each tube.
- For the tube No.6,pour off the 0.2ml
- Hence, the dilutions will range from 1:2 to 1:64.
- Take each dilution or titration and do a qualitative test as indicated above in order to determine the titer
Multiplication of the dilution factor by 6 mg/L will yield the approximate level of CRP present. The following table is only shown as an example for the determination of CRP concentration in specimen. Actual specimen will have CRP concentrations higher or lower than the levels indicated in this table.
CRP Normal range
A normal crp levels in blood or crp values are below 3.0 mg/L. Keep in mind that this normal reference range often varies between laboratoties. A high-sensitivity CRP test or elevated crp levels can detect levels below 10.0 mg/L
LIMITATIONS OF CRP TEST PROCEDURE
- The results of this test SHOULD NOT be used as a single diagnostic tool to make a clinical diagnosis. Instead, the test results must be evaluated together with other clinical findings and observed symptoms to aid in the final diagnosis.
- This test is designed to be performed by hand rotation. The use of a mechanical rotator could yield false positive/negative results.
- The strength of the agglutination reaction is not indicative of the CRP concentration.
- Weak reactions may occur with slightly elevated or markedly elevated concentrations
- A prozone phenomenon (antigen excess) may cause false negatives. It is advisable, therefore, to check all negative sera by retesting at a 1:10 dilution. Reaction times longer than specified may produce apparent false reactions due to a drying effect.
- Strongly lipemic or contaminated sera can cause false positive reactions.
- Only serum should be used in this test.
- Specimens containing Rheumatoid factor (RF) should not be used. The presence of RF can be confirmed by using commercially available RF Tests (e.g. Latex Tests). The presence of RF (usually >20 IU/ml) may lead to false