AnaSpec is pleased to announce
the release of a recombinant human BAFF protein. Naïve B-cells treated with
20-1000 ng/ml BAFF protein prepared by AnaSpec shows increased IgG secretion
compared to 2 leading commercial sources (Figure 1).
BAFF (BlyS, TALL-1, THANK,
TNFSF13B, zTNF4) belongs to the TNF superfamily of proteins and is expressed
by myeloid cells.1-4 It is a 285-amino acid long type II transmembrane
protein, with the soluble152-amino acid active region. 1-4 BAFF plays
a pivotal role in B-cells homeostasis including survival of immature B-cells,
transition of B-lymphocytes from T1 to T2 stage, and causes increase of antibodies
secretion by B-cells. 1-4
The sequence (Accession # NP_006564) corresponding to the extracellular
domain of human BAFF along with a 6x His tag was expressed in E. coli.
Recombinant human BAFF was purified from urea denatured bacterial lysate using
immobilized metal affinity chromatography (IMAC) and refolded into its active
state using a proprietary technique. The molecular weight of the recombinant
human BAFF is 17.2 kDa (Figure 2). Containing less than 0.1 EU of endotoxin
per 1 μg of protein as determined by LAL assay, the activity of BAFF was
determined by its ability to stimulate production of immunoglobulin molecules
such as IgG, IgA, and IgM in isolated mouse B-lymphocytes (Figure 1). Purified
recombinant human BAFF is supplied as sterile and frozen
condition at 0.1 mg/ml in 1 x PBS buffer with 5 mM DTT, pH=7.5.
Figure 1. Purified naïve B-lymphocytes were isolated
from mouse spleen, pre-treated with CpG ODN 1555, and stimulated with increasing
concentration of Human BAFF followed by IgG concentration measurement.”LPS+IL-4”
treated B-cells is the positive control and “0” the negative control.
Figure 2.Recombinant
human BAFF on 10-20% Tris-HCl SDS-PAGE. MWM=molecular weight marker, CL=cell
lysate before column coupling, FT=flow through after coupling, P=purified and
refolded recombinant human BAFF.
References:
- Batten, M. et. al. (2000) J Exp Med 192 (10): 1453-1465
- Parry, TJ. et. al. (2001) J Pharm Exp Ther 296 (2): 396-404
- Shan, X. et. al. (2006) Physiol Res 55: 301-307
- Katsenelson, N. et. al. (2007) Eur J Immunol 37: 1785-1795
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