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  3. Whitlow/218 Linker (E3U7Q) F(ab')<sub>2</sub> Fragment (Alexa Fluor® 647 Conjugate)
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Whitlow/218 Linker (E3U7Q) F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #47375

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Flow Cytometry Image 1: Whitlow/218 Linker (E3U7Q) F(ab')<sub>2</sub> Fragment (Alexa Fluor® 647 Conjugate)
Flow cytometric analysis of a mixed population containing live wild-type Jurkat cells and Jurkat cells engineered to express an scFv-based Anti-CD20 CAR containing a Whitlow/218 linker using Whitlow/218 Linker (E3U7Q) F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) (right) or concentration-matched Rabbit (DA1E) IgG XP® Isotype Control F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #93671 (left). Tag Blue fluorescent protein (TagBFP) is co-expressed with the CAR. Cell line was provided by the Lohmueller Lab, University of Pittsburgh.
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To Purchase # 47375

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 
  • Related Products

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

Supplied in PBS (pH 7.2), less than 0.1% sodium azide, and 2 mg/mL BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.

Protocol

Specificity / Sensitivity

Whitlow/218 Linker F(ab')2 fragment (Alexa Fluor® 647 Conjugate) recognizes exogenously expressed levels of scFv-based CARs containing a Whitlow/218 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 corresponding to residues of the Whitlow/218 linker peptide.

Background

The Whitlow/218 peptide is a commonly used linker that connects 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 Whitlow/218 linker affords the scFv with enhanced resistance to proteolysis and aggregation (1).

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.

限制使用

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For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is conditioned on the buyer using the purchased product solely in research conducted by the buyer, excluding contract research or any fee for service research, and the buyer must not (1) use this product or its components for (a) diagnostic, therapeutic or prophylactic purposes; (b) testing, analysis or screening services, or information in return for compensation on a per-test basis; or (c) manufacturing or quality assurance or quality control, and/or (2) sell or transfer this product or its components for resale, whether or not resold for use in research. For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008 USA or outlicensing@lifetech.com.
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