CAR-T Cell (G4S Linker) Transduction Efficiency Flow Cytometry Panel #18839
Product Information
Protocol
Product Description
The CAR-T Cell (G4S Linker) Transduction Efficiency Flow Cytometry Panel can be used to identify conventional human T cell subsets that have been engineered to express scFv-based CARs containing a G4S linker.
CD45 is a pan leukocyte marker. T cells are identified by expression of CD3. There are two major subsets of conventional T cells: helper T cells which express CD4, and cytotoxic T cells which express CD8. Monocytes are identified by the expression of CD14. CAR positive cells are identified by the expression of a G4S linker.
CD45 is a pan leukocyte marker. T cells are identified by expression of CD3. There are two major subsets of conventional T cells: helper T cells which express CD4, and cytotoxic T cells which express CD8. Monocytes are identified by the expression of CD14. CAR positive cells are identified by the expression of a G4S linker.
Product Usage Information
All antibodies in this kit are compatible with the Flow Cytometry, Live Cell Protocol for Directly Conjugated Antibodies and can be used in a single staining mix. After antibody staining and prior to acquisition on a flow cytometer, we recommend adding a membrane impermeable viability dye such as Propidium Iodide or 7-AAD to enable identification and exclusion of dead cells from the analysis.
For individual product dilutions, refer to panel product datasheet or individual product pages.
Gating strategy for identifying viable, CAR positive T cell subsets: Gate on live cells using Propidium Iodide or 7-AAD as a histogram or scatterplot with forward scatter (FSC). Use a gate set on side scatter (SSC) vs. FSC to exclude cell debris. Singlets may be identified by setting a gate on FSC-H vs. FSC-A. Next, use a plot of CD45 vs. CD3 to identify CD45+ leukocytes, and CD3+ and CD3- subpopulations. Observe G4S Linker vs. CD3 expression and set a gate on CD3+G4S Linker+ cells, which are CAR+ T cells. Observe CD4 vs. CD8 expression within the gated population to identify CD4+ and CD8+ T cells that are CAR+. Similarly, observe G4S Linker vs. CD14 expression within the CD3- gate to identify contaminating monocytes.
For individual product dilutions, refer to panel product datasheet or individual product pages.
Gating strategy for identifying viable, CAR positive T cell subsets: Gate on live cells using Propidium Iodide or 7-AAD as a histogram or scatterplot with forward scatter (FSC). Use a gate set on side scatter (SSC) vs. FSC to exclude cell debris. Singlets may be identified by setting a gate on FSC-H vs. FSC-A. Next, use a plot of CD45 vs. CD3 to identify CD45+ leukocytes, and CD3+ and CD3- subpopulations. Observe G4S Linker vs. CD3 expression and set a gate on CD3+G4S Linker+ cells, which are CAR+ T cells. Observe CD4 vs. CD8 expression within the gated population to identify CD4+ and CD8+ T cells that are CAR+. Similarly, observe G4S Linker vs. CD14 expression within the CD3- gate to identify contaminating monocytes.
Storage
CD3 (UCHT1) Mouse mAb (PE-Cy7® Conjugate), CD4 (RPA-T4) Mouse mAb (FITC Conjugate), CD8α (RPA-T8) Mouse mAb (PE Conjugate), CD45 (HI30) Mouse mAb (redFluor 710 Conjugate), and CD14 (61D3) Mouse mAb (violetFluor 450 Conjugate) are supplied in 10 mM NaH2PO4, 150 mM NaCl, 0.09% NaN3, 0.1% gelatin, pH 7.2. G4S Linker (E7O2V) Rabbit mAb (Alexa Fluor® 647 Conjugate) is supplied in PBS (pH 7.2), less than 0.1% sodium azide, and 2 mg/mL BSA. Store at 4ºC. Do not aliquot the antibodies. Protect from light. Do not freeze.
All components in this kit are stable in accordance with the date printed on the outer packaging label when stored at the recommended temperature. Please refer to product labels, datasheets, or web pages for specific “Best By” dates for each individual component.
All components in this kit are stable in accordance with the date printed on the outer packaging label when stored at the recommended temperature. Please refer to product labels, datasheets, or web pages for specific “Best By” dates for each individual component.
Specificity / Sensitivity
Each of the lineage marker antibodies in the CAR-T Cell (G4S Linker) Transduction Efficiency Flow Cytometry Panel detects endogenous levels of its target protein. CD3 (UCHT1) Mouse mAb (PE-Cy7® Conjugate) detects an epitope within the extracellular domain of CD3ε protein. CD4 (RPA-T4) Mouse mAb (FITC Conjugate ), CD8α (RPA-T8) Mouse mAb (PE Conjugate), CD45 (HI30) Mouse mAb (redFluor 710 Conjugate), and CD14 (61D3) Mouse mAb (violetFluor 450 Conjugate) detect epitopes within the extracellular domains. G4S Linker (E7O2V) Rabbit mAb (Alexa Fluor® 647 Conjugate) recognizes exogenously expressed levels of scFv-based CARs containing a G4S linker.
Species Reactivity:
Human
Source / Purification
Monoclonal antibodies were purified from tissue culture supernatant via affinity chromatography. The purified antibodies were conjugated under optimal conditions, with unreacted dye removed from the preparation.
限制使用
除非 CST 的合法授书代表以书面形式书行明确同意，否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件，除非书 CST 的合法授书代表以书面形式书独接受， 否书均被拒书，并且无效。
专品专有“专供研究使用”的专专或专似的专专声明， 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的， 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专，且专用于研专用途。将专品用于专断、专防或治专目的， 或专专售（专独或作专专成）或其他商专目的而专专专品，均需要 CST 的专独专可。客专：(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品，或使用专品制造任何商专专品，(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专，或使用专品开专任何与 CST 的专品或服专专争的专品或服专， (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专，(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品， (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专
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.
Cy and CyDye are registered trademarks of GE Healthcare.
All other trademarks are the property of their respective owners. Visit our
Trademark Information page.