HCG beta ELISA 试剂盒

This kit is for use with serum samples without additives only.

1. All reagent should be brought to room temperature (18-22 °C) before use.
   2. Reconstitute each lyophilized standard with 0.4 mL distilled water. Allow the reconstituted material to stand for at least 20 minutes. Reconstituted standards should be stored sealed at 2-8 °C, and it will be stable for at least two weeks at that conditions.
   3. Dilute 1 volume of Wash Buffer (50x) with 49 volumes of distilled water. For example, Dilute 15 mL of Wash Buffer Concentrate (50x) into distilled water to prepare 750 mL of washing buffer(1x). Mix well before use.

1. Secure the desired number of coated wells in the holder.
   2. Dispense 50 mL of standard, specimens, and controls into appropriate wells.
   3. Dispense 100 mL of Zero Buffer into each well.
   4. Thoroughly mix for 10 seconds. It is very important to have completed mixing in this setup.
   5. Incubate at 37 °C for 30 minutes.
   6. Remove the incubation mixture by flicking plate content into a sink.
   7. Rinse and empty the microtiter plate 4 times with washing buffer(1X) and 1 time with distilled water.
   8. Strike the wells sharply onto absorbent paper or paper towels to remov all residual water droplets.
   9. Dispense 150 mL of Enzyme Conjugate Reagent into each well. Gently mix for 5 seconds.
   10. Incubate at 37 °C for 30 minutes. Remove the incubation mixture by flicking plate contents into sink.
   11. Rinse and empty the microtiter plate 4 times with washing buffer(1X and 1 time with distilled water.
   12. Strike the wells sharply onto absorbent paper or paper towels to remove all residual water droplets.
   13. Dispense 100 mL TMB solution into each well. Gently mix for 5 seconds.
   14. Incubate at room temperature in the dark for 20 minutes.
   15. Stop the reaction by adding 100 mL of Stop Solution to each well.
   16. Gently mix for 4 till approx. 30 seconds. It is very important to make sure that the blue color changes to yellow color completely.
   17. Read optical density at 450 nm with a microtiter plate reader within 30 minutes.
   Important Note: The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbance reading.
   Limitations and Precaution: The free beta hCG-subunit quantitative assay is designed for in vitro use only. The components in this kit are intended for use as an integral unit. The components of different lots should not be mixed.

The absorbance of this quantitative assay is to 50 mIU/mL of beta-hCG. It is recommended that samples falling above the range should be diluted with sample diluent (Zero Buffer) to absorbance that within the standard curve range. Calculation of results Calculate the mean absorbance value (A450) for each set of reference standards, specimens, controls and patient samples. Constructed a standard curve by plotting the mean absorbance obtained from each reference standard against its concentration in ng/mL on log-log graph paper, with absorbance values on the vertical or Y axis and concentrations on the horizontal or X axis. Use the mean absorbance values for each specimen to determine the corresponding concentration of beta-hCG in mIU/mL from the standard curve. Example of standard curve Results of typical standard run with optical density reading at 450 nm shown in the Y axis against hCG concentrations shown in the X axis. This standard curve is for the purpose of illustration only, and should not be used to calculate unknowns. Each user should obtain his or her own data and standard curve.
Expected Values and Indications for Quantitative Free beta-hCG Assay:
1. In early pregnancy, free beta-hCG concentration was found 14 till approx. 80 mIU/mL. The free beta-hCG /intact hCG ratio was found 0.44 till approx. '0.48 percents. After 6 to 7 weeks the free beta-hCG and the ratio value declined. During the second and third trimester, a constant ratio was observed about 1 percent.
2. Serum samples from 40 normal subjects were assayed, in this population, 99 % of the values were less than 0.4 mIU/mL.