ATP5J ELISA Kit (Human)

Intra-assay Precision (Precision within an assay) : 3 samples with low, middle and high level Coupling Factor 6 (CF6) were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays) : 3 samples with low, middle and high level Coupling Factor 6 (CF6) were tested on 3 different plates, 8 replicates in each plate. CV (%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

ATP synthase subunit h; ATP synthase, H+ transporting, mitochondrial F0 complex, subunit F6; ATP synthase, H+ transporting, mitochondrial Fo complex subunit F6; ATP synthase-coupling factor 6, mitochondrial; ATP5; ATP5A; ATP5J; ATPase subunit F6; ATPM; CF6; coupling factor 6; F6; mitochondrial ATP synthase, coupling factor 6; mitochondrial ATP synthase, subunit F6; mitochondrial ATPase coupling factor 6; proliferation-inducing protein 36.

Mitochondrial membrane ATP synthase (F (1) F (0) ATP synthase or Complex V) produces ATP from ADP in the presence of a proton gradient across the membrane which is generated by electron transport complexes of the respiratory chain. F-type ATPases consist of two structural domains, F (1) - containing the extramembraneous catalytic core and F (0) - containing the membrane proton channel, linked together by a central stalk and a peripheral stalk. During catalysis, ATP synthesis in the catalytic domain of F (1) is coupled via a rotary mechanism of the central stalk subunits to proton translocation. Part of the complex F (0) domain and the peripheric stalk, which acts as a stator to hold the catalytic alpha (3) beta (3) subcomplex and subunit a/ATP6 static relative to the rotary elements. Also involved in the restoration of oligomycin-sensitive ATPase activity to depleted F1-F0 complexes.

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 Coupling Factor 6 (CF6) . Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Coupling Factor 6 (CF6) . 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 Coupling Factor 6 (CF6), 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 Coupling Factor 6 (CF6) 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 (%) Matrix Recovery range (%) Average (%) serum (n=5) 94-105 98 EDTA plasma (n=5) 80-90 86 heparin plasma (n=5) 82-91 86 Matrix Recovery Range (%) Average (%) Matrix Recovery Range (%) Average (%) Matrix Recovery range (%) Average (%) Matrix Recovery range (%) Average (%) serum (n=5) 94-105 98 serum (n=5) 94-105 98 EDTA plasma (n=5) 80-90 86 EDTA plasma (n=5) 80-90 86 heparin plasma (n=5) 82-91 86 heparin plasma (n=5) 82-91 86

Component Amount Anti-ATP5J Microplate 96 Wells (12 x 8 Well Strips) ATP5J Lyophilized Standard 2 x 10 ng 100X Biotinylated ATP5J 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-ATP5J Microplate 96 Wells (12 x 8 Well Strips) Anti-ATP5J Microplate 96 Wells (12 x 8 Well Strips) ATP5J Lyophilized Standard 2 x 10 ng ATP5J Lyophilized Standard 2 x 10 ng 100X Biotinylated ATP5J Detector Antibody 120 uL 100X Biotinylated ATP5J 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 Coupling Factor 6 (CF6) . No significant cross-reactivity or interference between Coupling Factor 6 (CF6) and analogues was observed.