Procollagen type 1 N-terminal propeptide (P1NP) is the N-terminal extension of procollagen type 1 removed proteolytically during collagen formation1. P1NP is a bone formation biomarker usually secreted into blood stream as unstable trimeric structure which subsequently undergo thermal degradation to form a stable monomeric structure2.

This assay is a quantitative sandwich ELISA. The immunoplate is pre-coated with a monoclonal antibody specific for human P1NP. Standards and samples are pipetted into the wells and any human P1NP present is bound by the immobilized antibody. After washing away any unbound substances, a biotin labelled polyclonal antibody specific for human P1NP 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 P1NP 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 P1NP, the unknown sample concentration can be interpolated from a reference curve included in each assay.

A. Typical representation of standard curve

The following standard curve is provided for demonstration only. A standard curve should be generated for each set of sample assay. 

Human  P1NP (pg/ml)	Absorbance (450 nm)	Blanked Absorbance
0	0.094	0
62.5	0.133	0.039
125	0.187	0.093
250	0.293	0.199
500	0.474	0.38
1000	0.812	0.718
2000	1.317	1.223
4000	2.096	2.002

 

B. Sensitivity

The lowest level of human PINP that can be detected by this assay is 62.5 pg/mL.

 

C. Precision

Intra-assay Precision (Precision within an assay) C.V. <3.3%.

Inter-assay Precision (Precision between assays) C.V. <5.1%.