Anti-Cytochrome P450 3A4/CYP3A4 抗体 [EPR6202]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human liver tissue sections labeling Cytochrome P450 3A4/CYP3A4 with purified ab124921 at 1/400 dilution (0.35 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

ab124921 (unpurified), at a 1/100 dilution, staining Cytochrome P450 3A4/CYP3A4 in paraffin embedded Human liver tissue by Immunohistochemistry.

• IP

Unknown

Immunoprecipitation - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

ab124921 (purified) at 1/20 dilution (1ug) immunoprecipitating Cytochrome P450 3A4/CYP3A4 in Human liver lysates.
Lane 1 : Human liver lysates 10ug
Lane 2 (+) : ab124921 & Human liver lysates
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab124921 in Human liver lysates
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was at 1/1000 dilution.
Blocking and diluting buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (ab124921)

Predicted band size: 57 kDa

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• WB

Unknown

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

All lanes:

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (ab124921) at 1/10000 dilution

All lanes:

Human fetal liver lysate at 10 µg

Secondary

All lanes:

HRP labelled goat anti rabbit at 1/2000 dilution

Predicted band size: 57 kDa

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• WB

Unknown

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

HepG2 cells were incubated at 37°C for 48h with vehicle control (0 μM) and different concentrations of nicardipine hydrochloride (ab120531) in DMSO. Increased expression of Cytochrome P450 3A4/CYP3A4 (ab124921, unpurified) correlates with an increase in nicardipine hydrochloride concentration, as described in literature.

Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 10μg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 5% BSA before being incubated with ab124921 (unpurified) at 1/10000 dilution and ab9484 at 1 μg /ml overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP (ab97051) at 1/10000 dilution and visualised using ECL development solution.

All lanes:

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (ab124921)

Predicted band size: 57 kDa

true

Exposure time: 20min

• WB

Lab

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

This antibody fails to detect BOI in normal HepG2 cells which is positive reported by PMID 26089940 and 25141173

All lanes:

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (ab124921) at 1/1000 dilution

Lane 1:

Human liver lysates at 20 µg

Lane 2:

HepG2 (Human hepatocellular carcinoma epithelial cell), Whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 57 kDa

Observed band size: 50 kDa

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• WB

Supplier Data

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

ab205606 was used as a tag loading control.

The detection of CYP3A4 recombinant protein or over -expressed protein with a C - terminal tag yielded a negative result because the immunogen of this product is located at the C - terminal of CYP3A4.

All lanes:

Western blot - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (ab124921) at 1/1000 dilution

Lane 1:

293T cells transfected with an empty vector containing a Flag tag whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human CYP3A4 expression vector containing a N-terminal Flag tag whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 50 kDa

false

Exposure time: 1s

• OI-RD Scanning

Supplier Data

OI-RD Scanning - Anti-Cytochrome P450 3A4/CYP3A4 antibody [EPR6202] (AB124921)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen) of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.