14-9918-82

antibody from Invitrogen Antibodies

Targeting: PAX5 BSAP

Provider product page for 14-9918-82

Western blot

Immunocytochemistry

Immunoprecipitation

Immunohistochemistry

Chromatin Immunoprecipitation

Other assay

Antibody data

Antibody Data
Antigen structure
References [5]
Comments [0]
Validations
Western blot [3]
Immunocytochemistry [1]
Chromatin Immunoprecipitation [2]
Other assay [4]

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Product number: 14-9918-82 - Provider product page
Provider: Invitrogen Antibodies
Product name: PAX5 Monoclonal Antibody (1H9), eBioscience™
Antibody type: Monoclonal
Antigen: Other
Description: Description: The monoclonal antibody 1H9 recognizes both mouse and human Pax5. Pax5, also known as BSAP (B cell specific activator protein), is a member of the paired box (pax) family of transcription factors. Pax5 is the only member of the pax family of transcription factors that is expressed in hematopoietic cells. During embryogenesis, Pax5 is transiently expressed in the brain of mice and in the mesencephalon and spinal cord of humans. Its expression is upregulated early in B cell development at the time of B cell commitment and is maintained throughout most subsequent stages. Suppression of Pax5 is essential for expression of Blimp-1 and the terminal differentiation of plasma cells. In the spleen, expression of Pax5 is higher in marginal zone B cells (B220+ CD21high CD23low) than in other B cells, especially the transition 1 stage (B220+ CD21- CD23-). In addition to its role in B cell development, Pax5 also affects VH-DJH heavy chain recombination as well as influencing the expression of many other B and non-B cell related proteins. Pax5 expression is correlated with many neoplasms. In diffuse large B cell lymphomas (DLBCL) and non-Hodgkin lymphomas Pax5 is often mutated while in B-cell ALL, expression levels are high. Additionally, translocation with elastin, IGH, ETV6, FOXP1, and EVI3 have been identified. eBioscience recommends that the Foxp3 Buffers (Product # 00-5521) be used for optimal results when using this antibody for intracellular staining and flow cytometric analysis. Applications Reported: This 1H9 antibody has been reported for use in immunoprecipitation, immunoblotting (WB), and immunohistology staining of frozen tissue sections. Applications Tested: This 1H9 antibody has been tested by western blot analysis. This can be used at less than or equal to 5 µg/mL. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest. Purity: Greater than 90%, as determined by SDS-PAGE. Aggregation: Less than 10%, as determined by HPLC. Filtration: 0.2 µm post-manufacturing filtered.
Reactivity: Human, Mouse
Host: Rat
Isotype: IgG
Antibody clone number: 1H9
Vial size: 100 µg
Concentration: 0.5 mg/mL
Storage: 4° C

PAX5 protein structure - 14-9918-82 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 7 PDK1 regulates Pax5 expression and cell-cycle progression in pre-B cells. To look at proliferation in vivo , PDK1 +/+ /Vav-Cre +ve and PDK1 fl/fl /Vav-Cre +ve mice were injected with 1 mg BrdU 3.25 h before being sacrificed. Bone marrow was then stained for 7-AAD and pre-B cell markers as described in Materials and methods. BrdU and 7-AAD uptake was determined by FACS in the pro-B and small pre-B cells ( A ). The levels of the proliferation marker Ki67 as well as cyclinD3 in ex vivo pre-B cells were also determined by FACS ( B ). To examine the expression of Pax5, IRF4, IRF8, Ikaros and Aiolos, total RNA was isolated from FACS-sorted pre-B cells and the mRNA levels for these genes determined by qPCR ( C ). Error bars represent the standard deviation of RNA from FACS-sorted pre-B cells of three independent pools of mice per genotype, P 0.05. The levels of Pax5, IRF4, IRF8, Ikaros and Aiolos protein ( D ) were determined by intracellular FACS staining of pro-B (CD19 +ve IgM -ve CD43 high B220 +ve ), large pre-B (FSC high CD19 +ve IgM -ve CD43 -ve B220 +ve ) or small pre-B (FSC low CD19 +ve IgM -ve CD43 -ve B220 +ve ). Results are representative of three mice per genotype.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 8 Pax5 and Bcl2A1 induce B-cell differentiation in the absence of PDK1. The levels of apoptosis in ex vivo PDK1 +/+ Vav-Cre +ve and PDK1 fl/fl Vav-Cre +ve cells were determined by annexin V staining of pro-B (CD19 +ve IgM -ve CD43 high B220 +ve ), large pre-B (FSC high CD19 +ve IgM -ve CD43 -ve B220 +ve ) or small pre-B (FSC low CD19 +ve IgM -ve CD43 -ve B220 +ve ) cells. Results are representative of three mice ( A ). qPCR was used to determine the mRNA levels of Mcl-1 and Bcl2A1 in FACS-sorted pre-B cells ( n =3) from PDK1 +/+ Vav-Cre +ve and PDK1 fl/fl Vav-Cre +ve mice ( B ). To determine if Pax5 and Bcl2A1 were involved in mediating PDK1 function in B cells, PDK1 fl/fl Vav-Cre +ve FACS-sorted CD19 +ve IgM -ve CD43 high B220 +ve DAPI -ve pro-B cells were infected with the indicated combinations of retroviruses for GFP, mCherry, GFP-Pax5 or mCherry-Bcl2A1 as described in Materials and methods. The percentages of live cells on day 6 in culture were determined based on forward and side scatter ( C , upper panels). BCR expression was then determined by surface staining for IgM and IgD in the GFP and/or mCherry-positive cells ( C , lower panels). Quantification of the % of live cells in ( C ) ( n =3) is shown in ( D ) while qualification of the % IgM/IgD positive gates for the indicated proteins in shown in ( E ). To analyse the effect on the cell cycle, PDK1 knockout pro-B cells were isolated and transfected with retroviruses for GFP-Pax5 and mCherry-Bcl2A1. After 6 day