G4S Linker (E7O2V) F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #42709
- F
Supporting Data
| REACTIVITY | All |
| SENSITIVITY | Transfected Only |
| MW (kDa) | |
| Source/Isotype | Rabbit IgG |
Application Key:
- F-Flow Cytometry
Species Cross-Reactivity Key:
- All-All Species Expected
Product Information
Product Description
This Cell Signaling Technology® F(ab’)2 fragment lacks an Fc domain, and will not exhibit reactivity with human Fc receptors. This eliminates the need for Fc blocking and reduces false positives in assays containing human monocytes or other cells with high expression of Fc receptors.
The F(ab')2 fragment is generated by enzymatic digestion of the full-length IgG clone coupled with chromatography to remove both the Fc fragment and the intact IgG. The F(ab’)2 fragment is conjugated to Alexa Fluor® 647 fluorescent dye under optimal conditions.
Product Usage Information
| Application | Dilution |
|---|---|
| Flow Cytometry (Live) | 1:50 |
Storage
Protocol
Specificity / Sensitivity
G4S Linker (E7O2V) F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) recognizes exogenously expressed levels of scFv-based CARs containing a G4S linker. This F(ab')2 fragment contains no Fc domain and will not exhibit cross-reactivity with Fc receptors.
Species Reactivity:
All Species Expected
Source / Purification
Monoclonal antibody is produced by immunizing animals with a synthetic peptide containing three Gly4Ser repeats.
Background
The poly-Glycine-Serine (G4S) linker is a type of flexible, unstructured synthetic peptide linker sequence often leveraged to connect the variable heavy (VH) domain and variable light (VL) domain of single-chain variable fragments (scFvs) and chimeric antigen receptors (CARs) that utilize an extracellular domain scFv for target antigen recognition. The linker itself consists of a core pentapeptide sequence, Gly-Gly-Gly-Gly-Ser, that is repeated and commonly found as either a 15-mer (G4S)3 or 20-mer (G4S)4 within scFv-based CARs and scFv fragments. The linker sequence length plays a role in controlling scFv stability and the noncovalent association between the VH and VL domains (1,2).
Human Fc receptors exhibit strong cross-reactivity with rabbit IgG, and are expressed at high levels on cells of myeloid lineage (e.g., monocytes and myeloid-derived cell lines). Fluorescent anti-linker F(ab')2 fragment conjugates are ideal for use in flow cytometry assays on cells containing Fc receptors, and eliminate the need for an Fc blocking step prior to antibody incubation.
Cell Signaling Technology employs strict quality control procedures to ensure that each conjugate provides optimal specificity and fluorescence.
Limited Uses
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