Tissue Factor Pathway Inhibitor ELISA
The Tissue Factor Pathway Inhibitor ELISA is For Research Use Only
Size: 12×8 wells
Sensitivity: 2.02 ng/ml
Standard Range: 30 ng/ml – 480 ng/ml
Incubation Time: 70 minutes
Sample Type: Serum, cell culture supernatant and plasma
Sample Size: 40 ul
Alternative Names: TFPI
The method employs sandwich ELISA technique. Monoclonal antibodies are pre-coated onto microwells. Samples and standards are pipetted into microwells and Human Tissue Factor pathway Inhibitor, TFPI present in the sample are bound by the antibodies. Biotin labeled antibody is added and followed by HRP (horseradish peroxidase) conjugated secondary antibody is pipetted and incubated to form a complex. After washing microwells in order to remove any non-specific binding, the ready to use substrate solution (TMB) is added to microwells and color develops proportionally to the amount of Human Tissue Factor pathway Inhibitor, TFPI in the sample. Color development is then stopped by addition of stop solution. Absorbance is measured at 450 nm.
Sample Preparation and Storage:
Specimens should be clear and non-hemolyzed. Samples should be run at a number of dilutions to ensure accurate quantitation.
1. Extract as soon as possible after specimen collection as per relevant procedure. The samples should be tested as soon as possible after the extraction. Alternately the extracted samples can be kept in -20°C. Avoid repeated freeze-thaw cycles.
2. Serum- Coagulate at room temperature for 10-20 minutes; centrifuge for 20-min at 2000-3000 rpm. Remove the supernatant. If precipitation appears, recentrifuge.
3. Plasma- Use EDTA or citrate plasma as an anticoagulant, mix for 10-20 minutes; centrifuge for 15-min at 2000-3000 rpm. Remove the supernatant carefully. If precipitation appears, recentrifuge.
4. Cell Culture Supernatant- Collect sample in a sterile container. Centrifuge for 20-mins at 2000-3000 rpm. Remove the supernatant carefully. When examining the components within the cell, dilute cell suspension with PBS (pH 7.2-7.4), if cell concentration is greater than 1 million/ml. Damage the cells by repeated freeze-thaw cycles to release intracellular components. Centrifuge for 20-min at 2000-3000 rpm. If precipitation appears, centrifuge again.
Note: Grossly hemolyzed samples are not suitable for use in this assay.