This CLIA kit can be applied to the in vitro quantitative determination of Human CASP3 concentrations in serum, plasma and other biological fluids.
This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Human CASP3. Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human CASP3 and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human CASP3, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Human CASP3. The concentration of Human CASP3 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
|Detection range||78.13-5000 pg/mL|
|Sample type||Serum, plasma and other biological fluids|
|Repeatability||CV < 15%|
This kit recognizes natural and some recombinant Human CASP3. No significant cross-reactivity or interference between Human CASP3 and analogues was observed.
For convenience in result calculation, absorbance can be used as ordinate and standard concentrations as abscissa. The standard curve provided in the manual is only for reference, and experimenters should draw the standard curve based on data of themselves.
Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Human CASP3 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level Human CASP3 were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||11.32||7.09||11.23||11.32||7.09||11.23|
The recovery of Human CASP3 spiked to three different levels in samples throughout the range of the assay in various matrices was evaluated.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||87-101||94|
|Cell culture media (n=5)||98-110||104|
Samples were spiked with high concentrations of Human CASP3 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
|Serum (n=5)||EDTA plasma (n=5)||Cell culture media (n=5)|
The unopened kit can be stored at 4 ℃ for 1 week. If the kit is not used within 1 week, store the items separately according to the following conditions since the kit is received.
|Micro CLIA Plate||8 wells ×12 strips||-20℃, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100×)||1 vial, 120 μL|
|Concentrated HRP Conjugate (100×)||1 vial, 120 μL||-20℃(shading light), 6 months|
|Reference Standard & Sample Diluent||1 vial, 20 mL||4℃, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25×)||1 vial, 30 mL|
|Substrate Reagent A||1 vial, 5 mL||4℃ (shading light)|
|Substrate Reagent B||1 vial, 5 mL|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
All reagent bottle cap must be tighten to prevent evaporation and microbial pollution.
The volume of reagents in partial shipments is a little more than the volume marked on the label, please use in measuring instead of directly pouring.
1.Add 100 μL of standard or sample to each well. Incubate for 90 min at 37℃.
2.Remove the liquid.
3.Add 100 μL Biotinylated Detection Ab. Incubate 1 hour at 37℃. Aspirate and wash 3 times.
4.Add 100 μL HRP Conjugate. Incubate 30 min at 37℃. Aspirate and wash 5 times.
5.Add 100 μL Substrate Mixture Solution. Incubate for 5 min at 37℃.
6.Fluorescence appeared. Measure the RLU value with the Chemiluminescence immunoassay analyzer. Calculation of the results.