BPI ELISA Kit (Mouse)

Intra-assay Precision (Precision within an assay) : 3 samples with low, middle and high level Bactericidal/Permeability Increasing Protein (BPI) were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays) : 3 samples with low, middle and high level Bactericidal/Permeability Increasing Protein (BPI) were tested on 3 different plates, 8 replicates in each plate. CV (%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

The cytotoxic action of BPI is limited to many species of Gram-negative bacteria; this specificity may be explained by a strong affinity of the very basic N-terminal half for the negatively charged lipopolysaccharides that are unique to the Gram-negative bacterial outer envelope.

The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to Bactericidal/Permeability Increasing Protein (BPI) . Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Bactericidal/Permeability Increasing Protein (BPI) . Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain Bactericidal/Permeability Increasing Protein (BPI), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm +/- 10nm. The concentration of Bactericidal/Permeability Increasing Protein (BPI) in the samples is then determined by comparing the O.D. of the samples to the standard curve.

Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Reconstitution & Storage Instructions Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Antibody Array (AA) Protocol

Matrix Recovery range (%) Average (%) Serum (n=5) 93-101 96 EDTA Plasma (n=5) 93-105 97 Heparin Plasma (n=5) 83-97 87 Matrix Recovery range (%) Average (%) Matrix Recovery range (%) Average (%) Serum (n=5) 93-101 96 Serum (n=5) 93-101 96 EDTA Plasma (n=5) 93-105 97 EDTA Plasma (n=5) 93-105 97 Heparin Plasma (n=5) 83-97 87 Heparin Plasma (n=5) 83-97 87

Component Amount Anti-BPI Microplate 96 Wells (12 x 8 Well strips) BPI Lyophilized Standard 2 100X Biotinylated BPI Detector Antibody 1 x 120 uL 100X Avidin-HRP Conjugate 1 x 120 uL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL Stop Solution 1 x 6 mL TMB Substrate 1 x 9 mL Component Amount Component Amount Anti-BPI Microplate 96 Wells (12 x 8 Well strips) Anti-BPI Microplate 96 Wells (12 x 8 Well strips) BPI Lyophilized Standard 2 BPI Lyophilized Standard 2 100X Biotinylated BPI Detector Antibody 1 x 120 uL 100X Biotinylated BPI Detector Antibody 1 x 120 uL 100X Avidin-HRP Conjugate 1 x 120 uL 100X Avidin-HRP Conjugate 1 x 120 uL Standard Diluent 1 x 20 mL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL 30X Wash Buffer 1 x 20 mL Stop Solution 1 x 6 mL Stop Solution 1 x 6 mL TMB Substrate 1 x 9 mL TMB Substrate 1 x 9 mL

This assay has high sensitivity and excellent specificity for detection of Bactericidal/Permeability Increasing Protein (BPI) . No significant cross-reactivity or interference between Bactericidal/Permeability Increasing Protein (BPI) and analogues was observed.