United Kingdom (UK)
Recombinant monoclonal antibody to Haemoglobin. Manufactured using AbAb’s Recombinant Platform with variable regions (i.e. specificity) from the hybridoma 2A1.
UniProt Accession Number of Target Protein: P01942
Alternative Name(s) of Target: Hemoglobin; Hb
Immunogen: Murine hybridomas were generated using spleen cells from aging (8–10 month old) NZB/W F1 animals. Hybridomas secreting antibodies reactive to murine hemoglobin were subcloned by limiting dilution and antibody isotypes determined using commercial reagents. Eight-week-old NZB/W F1 were immunized subcutaneously with 50 mg mHb antigen emulsified in Complete Fruend’s Adjuvant.
Specificity: 2A1 exhibited polyreactive recognition of peptide 100-119 from the alpha subunit and peptide 100-119 frmo the beta subunit. Moreover, 2A1 bound Hb and the Hb-haptoglobin complex with equal efficiency, implying different binding sites on Hb for haptoglobin and 2A1. 2A1 was also found to distinguish between Hb and metHb.
Application Notes: 2A1 has been used to bind to mHb in Western Blots and has been found to increase the secretion of TNF-alpha from endothelial cells when co-incubated with Hb, suggesting that 2A1 induces pro-inflammatory events (Bhatnagar 2011).
Antibody first published in:
Bhatnagar et al. Serum and organ-associated anti-hemoglobin humoralautoreactivity: Association with anti-Sm responses and inflammation Eur. J. Immunol. 2011. 41: 537–548 PMID:21268022
Note on publication:
Describes the presence of free hemoglobin in systemic lupus erythematosus and the investigation of anti-Hemoglobin autoimmune reactivity in patients.
Immunofluorescence staining of fixed mouse splenocytes with anti-Haemoglobin antibody 2A1 (Ab00956) Immunofluorescence analysis of paraformaldehyde fixed mouse splenocytes on Shi-fix™ coverslips, permeabilized with 0.15% Triton stained with the chimeric rabbit IgG version of 2A1 (Ab00956-23.0) at 10 µg/ml for 1h followed by Alexa Fluor® 488 secondary antibody (2 µg/ml), showing membrane staining. The nuclear stain is DAPI (blue). Panels show from left-right, top-bottom Ab00956-23.0, DAPI, merged channels and an isotype control. The isotype control was an unknown specificity antibody (Ab00178-1.1) followed by staining with Alexa Fluor® 488 secondary antibody.