Infinite Turbilatex CRP contain latex particles coated with human gammaglobulin which reacts with RF in the sample resulting in agglutination. Agglutination causes change in absorbance, measured at 650 nm (600 - 650 nm) & is proportional to the concentration of RF in the sample.
PREPARATION OF WORKING SOLUTION
R1 Tris buffer |
15 mmol/L |
45 ml |
R2 Latex particles coated with Gammaglobulin |
5 ml |
|
R3 : RF Calibrator (Value mentioned on vial label) |
2 ml |
SPECIMEN COLLECTION & PRESERVATION
Prepare the RF calibrator dilutions in normal saline as stated in the following table. Multiply the concentartion of the RF calibrator by the corresponding factor stated in the table to obtain the RF concentration of each dilution.
Calibrator dilution |
1 |
2 |
3 |
4 |
5 |
6 |
RF Calibrator(pt) Normal Saline (fii) |
100 |
10 90 |
25 75 |
50 50 |
75 25 |
100 - |
Factor |
0 |
0.1 |
0.25 |
0.5 |
0.75 |
1.0 |
Single point calibration (Linearity upto 100 IU/ml)
Prepare the RF calibrator dilution as : 30yl RF Calibrator + 70/zl normal saline. Multiply the concentartion of the RF calibrator by 0.30 to obtain the RF concentration of the diluted calibrator.
II. Manual Assay Procedure
Reagents |
Blank |
Calibrator |
Sample |
Working Solution Normal Saline Calibrator/Sample |
1.0 ml 7 yl |
1.0 ml 7 yI |
1.0 ml 7 yl |
Mix & read the absorbance after 2 minutes.
For multipoint calibration use all the 6 calibrator dilutions, while for single point calibration use only one diluted calibrator as stated above.
CALCULATION :
Calibration Curve
Plot the absorbance obtained for each calibrator dilution against the respective RF concentration. RF concentration in the sample is calculated by interpolation of its absorbance in the calibration curve.
Factor
Absorbance of Calibrator
RF (IU/ml) - Absorbance of Sample x Factor
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