SEMA4D ELISA Kit (Human)

Intra-assay Precision (Precision within an assay) : 3 samples with low, middle and high level Semaphorin 4D (SEMA4D) were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays) : 3 samples with low, middle and high level Semaphorin 4D (SEMA4D) were tested on 3 different plates, 8 replicates in each plate. CV (%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

A8; BB18; C9orf164; CD100; coll-4; COLL4; GR3; M-sema-G; sema domain, immunoglobulin domain (Ig), transmembrane domain (TM) and short cytoplasmic domain, (semaphorin) 4D; sema domain, immunoglobulin domain (Ig), transmembrane domain (TM) and short cytoplasmic domain, 4D; SEMAJ; semaphorin-4D.

Cell surface receptor for PLXN1B and PLXNB2 that plays an important role in cell-cell signaling. Promotes reorganization of the actin cytoskeleton and plays a role in axonal growth cone guidance in the developing central nervous system. Regulates dendrite and axon branching and morphogenesis. Promotes the migration of cerebellar granule cells and of endothelial cells. Plays a role in the immune system; induces B-cells to aggregate and improves their viability (in vitro) . Promotes signaling via SRC and PTK2B/PYK2, which then mediates activation of phosphatidylinositol 3-kinase and of the AKT1 signaling cascade. Interaction with PLXNB1 mediates activation of RHOA.

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 Semaphorin 4D (SEMA4D) . Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Semaphorin 4D (SEMA4D) . 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 Semaphorin 4D (SEMA4D), 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 Semaphorin 4D (SEMA4D) 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) 80-104 90 EDTA plasma (n=5) 94-102 99 heparin plasma (n=5) 78-101 83 Matrix Recovery Range (%) Average (%) Matrix Recovery Range (%) Average (%) serum (n=5) 80-104 90 serum (n=5) 80-104 90 EDTA plasma (n=5) 94-102 99 EDTA plasma (n=5) 94-102 99 heparin plasma (n=5) 78-101 83 heparin plasma (n=5) 78-101 83

Component Amount Anti-SEMA4D Microplate 96 Wells (12 x 8 Well Strips) SEMA4D Lyophilized Standard 2 x 100 ng 100X Biotinylated SEMA4D Detector Antibody 120 uL Avidin/HRP Conjugate 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 TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Component Amount Component Amount Anti-SEMA4D Microplate 96 Wells (12 x 8 Well Strips) Anti-SEMA4D Microplate 96 Wells (12 x 8 Well Strips) SEMA4D Lyophilized Standard 2 x 100 ng SEMA4D Lyophilized Standard 2 x 100 ng 100X Biotinylated SEMA4D Detector Antibody 120 uL 100X Biotinylated SEMA4D Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Avidin/HRP Conjugate 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 TMB Substrate 1 x 9 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Stop Solution 1 x 6 mL

This assay has high sensitivity and excellent specificity for detection of Semaphorin 4D (SEMA4D) . No significant cross-reactivity or interference between Semaphorin 4D (SEMA4D) and analogues was observed.