Human Total Serpin E1/PAI-1 Quantikine ELISA Kit

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DTSE100
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Human Total Serpin E1/PAI-1 Standard Curve
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Human Total Serpin E1/PAI-1 Quantikine ELISA Kit Summary

Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Assay Length
4.5 hours
Sample Type & Volume Required Per Well
Cell Culture Supernates (50 uL), Serum (10 uL), EDTA Plasma (10 uL), Heparin Plasma (10 uL), Citrate Plasma (10 uL)
Sensitivity
0.046 ng/mL
Assay Range
0.3 - 20 ng/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma, Citrate Plasma)
Specificity
Natural and recombinant human Total Serpin E1 (active, latent, uPA, tPA, and Vitronectin complexed Serpin E1).
Cross-reactivity
< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
Interference
No significant interference observed with available related molecules.

Product Summary

The Quantikine Human Total Serpin E1/PAI-1 Immunoassay is a 4.5 hour solid-phase ELISA designed to measure human Serpin E1 in cell culture supernates, serum, and plasma. It contains Sf 21-expressed recombinant human Serpin E1 and has been shown to accurately quantitate the recombinant factor free and in complex with uPA, tPA, or vitronectin. Results obtained using natural human Serpin E1 showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring human Serpin E1.

Precision

Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in twenty separate assays to assess inter-assay precision. Assays were performed by at least three technicians using two lots of components

Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma, Citrate Plasma

Intra-Assay Precision Inter-Assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 20 20 20
Mean 1.61 5.05 11.1 1.82 5.28 10
Standard Deviation 0.097 0.33 0.495 0.196 0.451 0.604
CV% 6 6.5 4.5 10.8 8.5 6

Recovery

The recovery of human Total Serpin E1 spiked to levels throughout the range of the assay in various matrices was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Media (n=4) 105 90-119
Citrate Plasma (n=4) 104 87-118
EDTA Plasma (n=4) 99 85-113
Heparin Plasma (n=4) 103 81-116
Serum (n=4) 104 93-116

Linearity

To assess the linearity of the assay, samples containing high concentrations of human Serpin E1 were serially diluted with Calibrator Diluent to produce samples with values within the dynamic range of the assay. All samples were diluted prior to assay.
Human Total Serpin E1/PAI-1 ELISA Linearity

Data Examples

Human Total Serpin E1/PAI-1 Standard Curve

Product Datasheets

Preparation and Storage

Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: Serpin E1/PAI-1

The human serpin superfamily consists of at least 35 members that target not only serine proteases, but also selected cysteine proteases and non-protease proteins. Serpins bind the protease active site resulting in a major conformational rearrangement that traps the enzyme in a covalent acyl-enzyme intermediate. As protease inhibitors, serpins have an array of functions including regulating blood clotting, the complement pathway, extracellular matrix remodeling, and cell motility. They are also involved in activities that extend beyond their ability to inhibit proteases. For instance, they may also regulate blood pressure, angiogenesis, or act as storage/transport proteins.

Long Name:
Plasminogen Activator Inhibitor
Entrez Gene IDs:
5054 (Human); 18787 (Mouse)
Alternate Names:
Endothelial plasminogen activator inhibitor; Nexin; PAI1; PAI-1; PAI1PAI-1; PAISerpin E1; PLANH1; PLANH1plasminogen activator inhibitor 1; serine (or cysteine) proteinase inhibitor, clade E (nexin, plasminogenactivator inhibitor type 1), member 1; Serpin E1; serpin peptidase inhibitor, clade E (nexin, plasminogen activator inhibitortype 1), member 1
⚠ WARNING: This product can expose you to chemicals including N,N-Dimethylforamide, which is known to the State of California to cause cancer. For more information, go to www.P65Warnings.ca.gov.

Assay Procedure

Refer to the product for complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
  1.   Prepare all reagents, standard dilutions, and samples as directed in the product insert.
  2.   Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

  3. 50 µL Assay Diluent
  4.   Add 50 µL of Assay Diluent to each well.

  5. 50 µL Standard, Control, or Sample
  6.   Add 50 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.
  7.   Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.

  8. 200 µL Conjugate
  9.   Add 200 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.
  10.   Aspirate and wash 4 times.

  11. 200 µL Substrate Solution
  12.   Add 200 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes. PROTECT FROM LIGHT.

  13. 50 µL Stop Solution
  14.   Add 50 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.

FAQs

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