PA5-28801

antibody from Invitrogen Antibodies

Targeting: ABCB1 ABC20, CD243, CLCS, GP170, MDR1, P-gp, PGY1

Provider product page for PA5-28801

Western blot

Other assay

Antibody data

Antibody Data
Antigen structure
References [2]
Comments [0]
Validations
Western blot [6]
Other assay [3]

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Product number: PA5-28801 - Provider product page
Provider: Invitrogen Antibodies
Product name: P-Glycoprotein Polyclonal Antibody
Antibody type: Polyclonal
Antigen: Synthetic peptide
Description: Recommended positive controls: HepG2, HepG2 membrane fraction extract, unboild HepG2, unboiled HepG2 membrane fraction extract. Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.
Reactivity: Human
Host: Rabbit
Isotype: IgG
Vial size: 100 µL
Concentration: 1.58 mg/mL
Storage: Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

ABCB1 protein structure - PA5-28801 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of P-Glycoprotein/MDR-1 using 20 µg of Hep3B lysate. Samples were loaded onto a 8% SDS-PAGE gel and probed with a P-Glycoprotein/MDR-1 polyclonal antibody (Product # PA5-28801) at a dilution of 1:2500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of P-Glycoprotein was performed by separating 30 µg of Boiled and unboiled HepG2 whole cell and membrane extracts by 5% SDS-PAGE. Proteins were transferred to a membrane and probed with a P-Glycoprotein Polyclonal Antibody (Product # PA5-28801) at a dilution of 1:500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of P-Glycoprotein was performed by separating 30 µg of Boiled and unboiled HepG2 whole cell and membrane extracts by 5% SDS-PAGE. Proteins were transferred to a membrane and probed with a P-Glycoprotein Polyclonal Antibody (Product # PA5-28801) at a dilution of 1:500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockout of ABCB1 was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042, Assay ID CRISPR634315_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of ABCB1 was performed by loading 30 µg of, Caco-2 Cas9 (Lane 1) andCaco-2 ABCB1 KO (Lane 2) membrane enriched extracts. The samples were electrophoresed using NuPAGE™ 3-8% Tris-Acetate Protein Gel (Product # EA0378BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with Anti-P-Glycoprotein Polyclonal Antibody (Product # PA5-28801, 1;3000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:5000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to ABCB1. An uncharacterized band was observed at ~270 kDa in all the samples.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of P-Glycoprotein was performed by separating 30 µg of Boiled and unboiled HepG2 whole cell and membrane extracts by 5% SDS-PAGE. Proteins were transferred to a membrane and probed with a P-Glycoprotein Polyclonal Antibody (Product # PA5-28801) at a dilution of 1:500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-P-Glycoprotein Polyclonal Antibody (Product # PA5-28801) and due to glycosylation, 160-250 kDa band corresponding to ABCB1 was observed across cell lines tested except MCF-7 which is reported to be negative. Whole cell extracts (30 µg lysate) of HepG2 (Lane 1), Caco-2 (Lane 2) and MCF-7 (Lane 3) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # LC2001) by XCell SureLock™ Mini-Cell and XCell II™ Blot Module (Product # EI0002). The blot was probed with the primary antibody (1:3000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 Met affects the expression of HIF-1alpha, MDR1/P-gp and MRP1. (A) Reverse transcription-polymerase chain reaction analysis of MDR1 and MRP1 expression in the different treatment groups. (B) Western blot analysis of HIF-1alpha, MDR1 and MRP1 in the different treatment groups. (C-E) Relative density of the western blotting bands demonstrated that levels of HIF-1alpha, P-gp and MRP1 decreased with increasing concentrations of Met in a dose-dependent manner, with a statistically significant difference between the treatments and the controls ( ** P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 5 FOLFOX induces CD44v6 expression, which is critically regulated by the WNT pathway stimulated by PN and/or IL17A. (A) , Confirmation of involvement of CD44v6 expression in regulating WNT3A production was assessed in PD-FR CAFs by examining the effects of blockade of CD44v6 using specific shRNAs on 10 4 SW480-FR/CAFs in the absence and presence of FOLFOX. The effects were evaluated by measuring secreted WNT3A in cultures by ELISA. (B) , Involvement of CD44v6 in regulating PN and IL17A induced WNT3A production was assessed in PD-FR CAFs by examining the effects of blockade of CD44v6 using specific shRNAs in the absence and presence of PN or IL17A. The effects were evaluated by measuring secreted WNT3A in cultures by ELISA. (C) , Schematic illustration of the CD44 gene is shown. Both constitutive (C) and variable (v) exons are represented. The PCR primers used to amplify CD44 variable (v) and standard (s) isoforms are shown as black arrows. The primers for both the v6 and standard isoforms (CD44s) predominantly generate one PCR product [C5v6 (v6) for CD44v6, and C5C7 for CD44s], whereas the primers for the v8 variants amplify two splice variants C5v6v7v8 (v6-v8) and C5v8 (v8). These PCR products are depicted in panels (D-G) experiments. The primers for both the v6 and standard isoforms predominantly generate one PCR product [c5v6 (v6)], whereas the primers for the v8 variants amplify two splice variants C5v6v7v8 (v6-v8), and C5v8 (v8). (D, E) , WNT3A shRNA, or CD44v6 sh

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Tissue specific knockdown of CD44v6 by pFabpl-Cre inhibits subcutaneous (SQ) tumor growth of implanted CICs plus CAFs from SW480-FR/SQ tumors and from PD-FR/SQ tumors. (A) , Timeline is shown for pSico-v6-shRNA/Nano particle treatment with and without pFabpl-Cre/Nano in xenograft tumors derived from CICs plus CAFs from SW480-FR/SQ tumors and PD-FR/SQ tumors. (B) , The dependence of tumors derived from CICs and CAFs on CD44v6 was evaluated in vivo. 2 x 104 FACS sorted CAM(+)CD44v6(+)CD133(+)ALDH1(+) CICs and 6 x 104 CAFs (EpCAM [-]/PDGFR-alpha were injected into mice. When tumors reached to approximately 0.3 cm3 in volume, treatment was initiated. Detailed treatment procedure is in Methods. Seven immunocompromised mice per group were used. Mice were weighed every other day, and the tumor weights were measured every week for 4 weeks. Representative tumors following sacrifice from three experiments are shown from mice treated as indicated in the schedule of treatment (A) . (C, D) , Kinetics of relative tumor weights with time are shown during in vivo SQ tumor growth at the indicated weeks that were generated by CICs plus CAFs from SW480-FR/SQ tumors (C) and from PD-FR/SQ tumors (D) injected (i.p.) with pSico-NT shRNA/Nano, pSico-v6 shRNA/Nano, and (pSico-v6 sh plus pFabpl-Cre)/Nano. Treatments were performed at weeks (wks) 2, 3, 4, and 5 and tumor growth was measured at wks 3, 4, 5, and 6 after treatments. (E) , Western blot analyses using CD44v6, MDR1 and beta-catenin antibodie