mAbs Against HUMAN Protein
PRDM1/Blimp-1
| STATUS: |  |
|---|---|
| CONTACT INFORMATION: | Monoclonal Antibodies Unit, Centro Nacional de Investigaciones Oncológicas |
| STATUS: | Validated |
| TYPE: | mouse anti human |
| CLONE NAME: | ROS195 |
| PROTEIN: | Human full length PRDM1 |
| PROTEIN WEB: | http://www.ncbi.nlm.nih.gov/omim/603423 |
| ANTIGEN USED: | GST-PRDM1 recombinant protein |
| FUSION PARTNER: | NS1/Ag4-1 (NS1) cells |
| ISOTYPE: | IgG1 |
| SPECIES REACTIVITY: | human and mouse |
| PREPARATION AND STORAGE: | Aliquot and store at 4C. Do not freeze |
| APP RECOMMENDED: | IHQ-paraffin, IF, WB, IP, Flow cytometry |
| APP NO TESTED: | IHQ-frozen |
Description
B lymphocyte-induced maturation protein-1 (Blimp-1) is a 98-kDa protein containing five Kruppel-type zinc fingers that confer sequence specific DNA binding. Based on studies in B cell lines, Blimp-1 has been postulated to be a master regulator of terminal B cell differentiation. In the BCL-1 lymphoma model of differentiation from a mature B cell to a plasma cell, ectopic expression of Blimp-1 is sufficient to cause terminal differentiation evidenced by loss of surface Ig, IgM secretion, expression of syndecan-1 on the cell surface, and cessation of cell division.
Publication describing antibody characterization/validation
Garcia JF, Roncador G, Garcia JF, Sanz AI, Maestre L, Lucas E, Montes-Moreno S, Fernandez Victoria R, Martinez-Torrecuadrara JL, Marafioti T, Mason DY, Piris MA. PRDM1/BLIMP-1 expression in multiple B and T-cell lymphoma. Haematologica. 2006. Apr; 91(4):467-74.
References
Liu YY, Leboeuf C, Shi JY, Li JM, Wang L, Shen Y, Garcia JF, Shen ZX, Chen Z, Janin A, Chen SJ, Zhao WL. Rituximab plus CHOP (R-CHOP) overcomes PRDM1-associated resistance to chemotherapy in patients with diffuse large B-cell lymphoma. Blood 2007. Jul1; 110(1): 339-44.
Zhao WL, Liu YY, Zhang QL, Wang L, Leboeuf C, Zhang YW, Ma J, Garcia JF, Song YP, Li JM, Shen ZX, Chen Z, Janin A, Chen SJ. PRDM1 is involved in chemoresistance of T-cell lymphoma and down-regulated by the proteasome inhibitor. Blood 2008. Apr 1;111(7): 3867-71.
Nie K, Gomez M, Landgraf P, Garcia JF, Liu Y, Tan LH, Chadburn A, Tuschl T, Knowles DM, Tam W. MicroRNA-mediated down-regulation of PRDM1/Blimp-1 in Hodgkin/Reed-Sternberg cells: a potential pathogenetic lesion in Hodgkin lymphomas. Am J Pathol. 2008. Jul; 173(1): 242-52.
Garcia JF, Roncador G, Garcia JF, Sanz AI, Maestre L, Lucas E, Montes-Moreno S, Fernandez Victoria R, Martinez-Torrecuadrara JL, Marafioti T, Mason DY, Piris MA. PRDM1/BLIMP-1 expression in multiple B and T-cell lymphoma. Haematologica. 2006. Apr; 91(4):467-74.
Applications
| IHC Techniques | Clone | Dilution | Antibody concentration | Antigen retrieval method | Visualization kit | +/- control | Protein localization | Positivity in other species | Protocol |
|---|---|---|---|---|---|---|---|---|---|
| Frozen tissue and cytospins | |||||||||
Recommended Result obtained is satisfactory. The reagent can be use in this application | ROS | 1:2 | supernatant | none | DAKO goat&mouse HRP | tonsil / muscle | nuclear | mouse | |
| Paraffin tissue | |||||||||
Recommended Result obtained is satisfactory. The reagent can be use in this application | ROS | 1:10 | supernatant | Tris-EDTA | Novolink Kit | tonsil / muscle | nuclear | mouse | Available |
| Immunofluorescence | |||||||||
Recommended Result obtained is satisfactory. The reagent can be use in this application | ROS | 1:4 | supernatant | tris-EDTA | Alexa goat&mouse 488 | tonsil / muscle | nuclear | mouse | Available |
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- ROS195 monoclonal antibody in transfected cells
- Nuclear staining on frozen cytospin preparations of transfected HEK293T/PRDM1 cells using antibody ROS195.
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- PRDM-1/Blimp-1 (ROS195) IHQ in human paraffin sections.
- In tonsil PRDM1 protein is strongly expressed by plasma cells present in germinal centres, and in subepithelial areas.
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- Triple immunostaining of PRDM1/Blimp-1 (ROS195) in reactive tonsil.
- PRDM1/Blimp-1 nuclear protein (blue) is expressed in plasma cells (VS38c, plasma cell marker, red) but absent in CD20 positive (B lymphocytes, green).
| WB Techniques | Clone | Dilution | Antibody concentration | +/- control | Expected MW | Observed Mw | Positivity in other species | Protocol |
|---|---|---|---|---|---|---|---|---|
| Western Blotting | ||||||||
Recommended Result obtained is satisfactory. The reagent can be use in this application | ROS | neat | supernatant | tonsil / muscle | 98kDa | 97kDa | not done | Available |
| Immunoprecipitation | ||||||||
Recommended Result obtained is satisfactory. The reagent can be use in this application | ROS | neat | supernatant | tonsil / muscle | 98kDa | 97kDa | not done | |
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- Biochemical characterization of ROS195 monoclonal antibody.
- Western blot analysis of PRDM1 expression in total protein extracts from lymphoma cell lines and human tonsil. Tubulin was used as loading control.
Lane 1 L-540 cell line (100ug) (+)
Lane 2 L-428 cell line (100ug) (+)
Lane 3 HDLM-2 cell line (100ug) (+)
Lane 4 KM-H2 cell line (100ug) (-)
Lane 5 L-1236 cell line (100ug) (-)
Lane 6 Human tonsil (100ug) (+)
Lane 7 RPMI-8226 cell line (100ug) (+)
Lane 8 Karpas-422 cell line (100ug) (+)
Lane 9 PBL (EBV) cell line (100ug) (-)
Lane 10 LP-1 cell line (100ug) (+)
Tubulin was used as loading control.
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- PRDM1/Blimp-1 (ROS195) immunoprecipitation.
- Immunoprecipitation of protein extracts from normal tonsil with the anti-PRDM1 monoclonal antibody (ROS195 lane) and the specific polyclonal antibody (pAb lane) followed by a Western blotting with ROS195 antibody, shows a ~97 kDa band.
| FLOW CYTOMETRY | Clone | Dilution | +/- control | Type of fluorocrom | Protocol |
|---|---|---|---|---|---|
Recommended Result obtained is satisfactory. The reagent can be use in this application | ROS | 50ul supernatant/one million cells/tube | OPM2 cell line / MOLT4 cell line |
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- ROS mAb by Flow cytometry
- RED: Negative cell line MOLT4 (T cell leukemia)
BLUE: Positive cell line OPM2 (multiple myeloma)
| OTHERS | Title | Description | Protocol |
|---|---|---|---|
Recommended Result obtained is satisfactory. The reagent can be use in this application | Epitope recognise by ROS mAb | ROS) monoclonal antibody recognize site between amino acids 176 and 307 and is able to recogne the two blimp-1 isoforms. |
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- Epitope recognised by ROS mAb
- ROS monoclonal antibody recognize site between amino acids 176 and 307 and is recognising the two blimp-1 isoforms. Work done by Dr K. Wright (Lee Moffitt Cancer Center)