MABSol® Biotinylated Human CD19, Fc Tag (CD9-H8259) is expressed from human HEK293 cells. It contains AA Pro 20 - Lys 291 (Accession # AAH06338). It is the biotinylated form of Human CD19 Protein, Fc Tag (Cat. No. CD9-H5259).
Predicted N-terminus: Pro 20
This protein carries a human IgG1 Fc fragment at the C-terminus. The protein has a calculated MW of 56.3 kDa. The protein migrates as 56-66 kDa on a SDS-PAGE gel under reducing (R) condition due to glycosylation.
The primary amines in the side chains of lysine residues and the N-terminus of the protein are conjugated with biotins using standard chemical labeling method. A standard biotin reagent (13.5 angstroms) is used in this product.
The biotin to protein ratio is 2.5-3.5 as determined by the HABA assay.
Less than 1.0 EU per μg by the LAL method.
>95% as determined by reduced SDS-PAGE.
Lyophilized from 0.22 μm filtered solution in PBS, pH7.4. Normally trehalose is added as protectant before lyophilization.
Contact us for customized product form or formulation.
Please see Certificate of Analysis for specific instructions.
For best performance, we strongly recommend you to follow the reconstitution protocol provided in the CoA.
For long term storage, the product should be stored at lyophilized state at -20°C or lower.
Please avoid repeated freeze-thaw cycles.
No activity loss was observed after storage at:
- 4-8°C for 12 months in lyophilized state;
- -70°C for 3 months under sterile conditions after reconstitution.
Biotinylated Human CD19, Fc Tag on SDS-PAGE under reducing (R) condition. The gel was stained overnight with Coomassie Blue. The purity of the protein is greater than 95%.
Immobilized Anti-Mouse FMC63 Mab at 2 μg/mL (100 μL/well) can bind Biotinylated Human CD19, Fc Tag (Cat. No. CD9-H8259) with a linear range of 0.039-0.625 μg/mL (QC tested).
Immobilized a series of concentration of Biotinylated Human CD19, Fc Tag (Cat. No. CD9-H8259) on streptavidin precoated (0.5 μg/well) plate, can bind FMC63 (Mouse IgG2a) 2 μg/mL (100 μL/well) with a linear range of 0.039-1.25 μg/mL (Routinely tested).
CAR阳性表达率检测（Evaluation of CAR expression ）
FACS Analysis of Anti-CD19 CAR Expression
293 cells were transfected with FCM63-scFv and RFP tag . 2x105 of the cells were first incubated with A. Biotinylated protein control. B. Recombinant biotinylated human CD19 (Cat. No. CD9-H8259, 10 μg/ml ) . C. Recombinant biotinylated human CD19 (Cat. No. CD9-H8259, 10 μg/ml) and FMC63(Mouse anti-CD19 antibody). FITC Streptavidin was used to analyse with FACS. RFP was used to evaluate CAR(FMC63-scFv) expression and FITC was used to evaluate the binding activity of recombinant biotinylated human CD19 (Cat. No. CD9-H8259) .
FACS Analysis of Anti-CD19 CAR Expression
Human T cells were lentivirally transduced with anti-CD19 CAR and cultured for 11 days. Eleven days post-transduction, 1e6 cells were stained for the expression of CD3 and anti-CD19 CAR with FITC anti-human CD3 antibody and biotinylated human CD19 (Cat. No. CD9-H8259) followed by PE-conjugated streptavidin, respectively. A. Non-transduced T cells were used as a control for gating of CAR expression. (Data are kindly provided by Beijing Bowei Huaen Medical Technology Co. Ltd.)
Authors: MacLeod DT, et al.
Journal: Mol Ther 2017
Application: Flow Cytometry
B-lymphocyte antigen CD19, is a single-pass type I membrane protein which contains two Ig-like C2-type (immunoglobulin-like) domains. CD19 is expressed on follicular dendritic cells and B cells. Upon activation, the cytoplasmic tail of CD19 becomes phosphorylated, which leads to binding by Src-family kinases and recruitment of PI-3 kinase. As on T cells, several surface molecules form the antigen receptor and form a complex on B lymphocytes. The (almost) B cell-specific CD19 phosphoglycoprotein is one of these molecules. The others are CD21 and CD81. These surface immunoglobulin (sIg)-associated molecules facilitate signal transduction. On living B cells, anti-immunoglobulin antibody mimicking exogenous antigen causes CD19 to bind to sIg and internalize with it. The reverse process has not been demonstrated, suggesting that formation of this receptor complex is antigen-induced. This molecular association has been confirmed by chemical studies. Mutations in CD19 are associated with severe immunodeficiency syndromes characterized by diminished antibody production. CD19 has been shown to interact with: CD81, CD82, Complement receptor 2, and VAV2.