PA5-29665

antibody from Invitrogen Antibodies

Targeting: GPI AMF, NLK

Antibody data

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Validation data

Product number: PA5-29665 - Provider product page
Provider: Invitrogen Antibodies
Product name: GPI Polyclonal Antibody
Antibody type: Polyclonal
Antigen: Recombinant protein fragment
Description: Recommended positive controls: H1299, Raji. Predicted reactivity: Rat (89%), Xenopus laevis (81%), Pig (92%), Rabbit (92%), Chicken (86%), Chimpanzee (99%), Bovine (90%). Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.
Reactivity: Human, Rat
Host: Rabbit
Isotype: IgG
Vial size: 100 µL
Concentration: 1 mg/mL
Storage: Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

GPI protein structure - PA5-29665 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of GPI was achieved by transfecting Hep G2 with GPI specific siRNAs (Silencer® select Product # s5982 ). Western blot analysis (Fig. a) was performed using whole cell extracts from the GPI knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Endothelin B Receptor Polyclonal Antibody (Product # PA5-29665, 1:5000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25µg/ml, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to GPI.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of Hep G2 (Lane 1), MCF7 (Lane 2), HeLa (Lane 3), Raji (Lane 4) and tissue extract of Rat Eye (Lane 5). The blot was probed with Anti-GPI Polyclonal Antibody (Product # PA5-29665, 1:5000 dilution) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/ml, 1:4000 dilution). A 55 kDa band corresponding to GPI was observed across cell lines and tissue tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of GPI was performed by separating 30 µg of various whole cell extracts by 7.5% SDS-PAGE. Proteins were transferred to a membrane and probed with a GPI Polyclonal Antibody (Product # PA5-29665) at a dilution of 1:2000 and a HRP-conjugated anti-rabbit IgG secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot using GPI Polyclonal Antibody (Product # PA5-29665). Sample (30 µg of whole cell lysate). Lane A: H1299. Lane B: Raji . 7.5% SDS PAGE. GPI Polyclonal Antibody (Product # PA5-29665) diluted at 1:10,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of GPI showing staining in the cytoplasm of HepG2 cells. HepG2 cells were fixed in ice-cold MeOH for 5 min and stained using a GPI polyclonal antibody (Product # PA5-29665) diluted at 1:500. Blue: Hoechst 33342 staining.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of GPI was performed using 70% confluent log phase HeLa cells. The cells were fixed with ice cold acetone for 5 minutes and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with GPI Polyclonal Antibody (Product # PA5-29665) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic and nuclear localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry-Immunofluorescence analysis of GPI was performed in HepG2 cells fixed in ice-cold MeOH for 5 min. Green: GPI Polyclonal Antibody (Product # PA5-29665) diluted at 1:500. Blue: Hoechst 33342 staining.