PA5-29580

antibody from Invitrogen Antibodies

Targeting: B2M

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Immunoprecipitation (Data presented on provider website)

Immunohistochemistry (Supportive data in Antibodypedia)

Other assay (Supportive data in Antibodypedia)

Antibody Data

Product number

PA5-29580

Provider

Invitrogen Antibodies

Product name

beta-2 Microglobulin Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Recombinant full-length protein

Description

Recommended positive controls: A431, mouse plasma, rat plasma, HeLa, B2M shRNA-transfected HeLa, marmoset rProtein. Predicted reactivity: Rhesus Monkey (92%), Chimpanzee (100%). Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.

Reactivity

Human, Mouse, Rat

Host

Rabbit

Isotype

IgG

Vial size

100 µL

Concentration

0.37 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of beta-2 Microglobulin was performed by separating 50 µg of mouse plasma by 15% SDS-PAGE. Proteins were transferred to a membrane and probed with a beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) at a dilution of 1:1500. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot analysis of beta-2 Microglobulin was performed by separating 50 µg of rat tissue extract by 15% SDS-PAGE. Proteins were transferred to a membrane and probed with a beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) at a dilution of 1:500.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockout of B2M was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042, Assay ID CRISPR983707_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of B2M was performed by loading 30 µg of HeLa Wild Type (Lane 1), HeLa Cas9 (Lane 2) andHeLa B2M KO (Lane 3) membrane enriched extracts. The samples were electrophoresed using NuPAGE™ Novex™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). 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-beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580, 1:1,000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4,000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to B2M.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot using beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580). Sample (30 µg of whole cell lysate). Lane A: A431 . 15% SDS PAGE. beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) diluted at 1:5,000.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of beta-2 Microglobulin was achieved by transfecting HeLa with beta-2 Microglobulin specific siRNAs (Silencer® select Product # s1854, s1853). Western blot analysis (Fig. a) was performed using Whole cell extracts from the beta-2 Microglobulin knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580, 1:500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 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 beta-2 Microglobulin.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) and a 12 kDa band corresponding to beta-2 Microglobulin was observed across in HeLa, THP-1 and U-937 but not in SH-SY5Y or NTERA-2 cell lines which are reported to express low levels of beta-2 microglobulin. Whole cell extracts (60 µg lysate) of HeLa (Lane 1), THP-1 (Lane 2), U-937 (Lane 3), SH-SY5Y (Lane 4) and NTERA-2 cl.D1 (Lane 5) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:500) 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 Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

beta-2 Microglobulin Polyclonal Antibody detects beta 2 Microglobulin protein by immunofluorescent analysis. Sample: HeLa cells were fixed in 4% paraformaldehyde at RT for 15 min. Green: beta 2 Microglobulin stained by beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) diluted at 1:500. Blue: Fluoroshield with DAPI .

Immunohistochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

beta-2 Microglobulin Polyclonal Antibody detects B2M protein at cytosol on Cal27 xenograft by immunohistochemical analysis. Sample: Paraffin-embedded Cal27 xenograft. Beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) dilution: 1:500. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

beta-2 Microglobulin Polyclonal Antibody detects beta 2 Microglobulin protein at cytoplasm in human endometrial cancer by immunohistochemical analysis. Sample: Paraffin-embedded human endometrial cancer. Beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) diluted at 1:500. Antigen Retrieval: Citrate buffer, pH 6.0, 15 min.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Beta 2 Microglobulin antibody detects secreted B2M protein by immunohistochemical analysis. Sample: Paraffin-embedded mouse spleen. B2M stained by Beta 2 Microglobulin antibody (Product # PA5-29580) diluted at 1:500. Antigen Retrieval: Citrate buffer, pH 6.0, 15 min.

Other assay

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

beta-2 Microglobulin Polyclonal Antibody immunoprecipitates B2M protein in IP experiments. IP samples: HeLa whole cell extract. A. 30 µg HeLa whole cell extract. B. Control with 4 µg of preimmune Rabbit IgG. C. Immunoprecipitation of B2M protein by 4 µg beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580). 15 % SDS-PAGE. The immunoprecipitated B2M protein was detected by beta-2 Microglobulin Polyclonal Antibody (Product # PA5-29580) diluted at 1:500.