Human Proinsulin ELISA Kit
★ Download MSDS ★
Assay range: 31.2 - 2000 pg/mL
Sample size: 50 uL
Size: 96 wells/kit
INTRODUCTION
Proinsulin is a polypeptide of 86 amino acids made in the beta cells of the pancreas and is the precursor molecule for insulin, most proinsulin is converted to insulin and C-peptide secreted into the blood. Increased circulating levels of proinsulin are found in patients with insulinomas, hypoglycemia and hyperinsulinema.
PRINCIPLE OF THE ASSAY
This assay is a quantitative sandwich ELISA. The immunoplate is pre-coated with a monoclonal antibody specific for human proinsulin. Standards and samples are pipetted into the wells and any human proinsulin present is bound by the immobilized antibody. After washing away any unbound substances, a biotin labelled monoclonal antibody specific for human proinsulin is added to the wells. After wash step to remove any unbound reagents, streptavidin-HRP conjugate (STP-HRP) is added. After the last wash step, an HRP substrate solution is added and colour develops in proportion to the amount of human proinsulin bound initially. The assay is stopped and the optical density of the wells determined using a microplate reader. Since the increases in absorbance are directly proportional to the amount of captured human proinsulin, the unknown sample concentration can be interpolated from a reference curve included in each assay.
ASSAY PERFORMANCE
A. Typical representation of standard curve
The following standard curve is provided for demonstration only. A standard curve should be generated for each assay.
Human proinsulin (pg/mL)
Absorbance
(450 nm)
Blanked Absorbance
0
0.096
0
31.2
0.122
0.026
62.5
0.154
0.058
125
0.225
0.129
250
0.354
0.258
500
0.616
0.52
1000
1.214
1.118
2000
2.168
2.072
B. Sensitivity
The lowest insulin level that can be measured by this assay is 31.2 pg/mL.
C. Precision
Intra-assay Precision (Precision within an assay) C.V. < 6.5%.
Inter-assay Precision (Precision between assays) C.V. <7.3%.
D. Recovery
The recovery of the assay was determined by adding various amounts insulin to a sample. The measured concentration of the spiked sample in the assay was compared to the expected concentration. The average recovery was 89.9%.
E. Linearity
98-113%