Each lot of this antibody is quality control tested by immunofluorescent staining with flow cytometric analysis and the oligomer sequence is confirmed by sequencing. TotalSeq?-C antibodies are compatible with 10x Genomics Chromium Single Cell Immune Profiling Solution.

To maximize performance, it is strongly recommended that the reagent be titrated for each application, and that you centrifuge the antibody dilution before adding to the cells at 14,000xg at 2 - 8°C for 10 minutes. Carefully pipette out the liquid avoiding the bottom of the tube and add to the cell suspension. For Proteogenomics analysis, the suggested starting amount of this reagent for titration is ≤ 1.0 ?g per million cells in 100 ?L volume. Refer to the corresponding TotalSeq? protocol for specific staining instructions.

The UC10-4B9 antibody can enhance T cell co-stimulation by blocking CTLA-4 interactions with the B7 co-receptors, favoring CD28 interactions. Additional reported applications (for the relevant formats) include: immunoprecipitation¹, in vitro stimulation, in vitro and in vivo blocking^1-4 of ligand binding, and as ELISA capture antibody⁵. To reduce non-specific binding to cells bearing Fc-receptors, pre-incubation of cells with anti-mouse CD16/CD32, clone 93 (Cat. No. 101301/101302), is recommended prior to immunofluorescent staining. For most successful immunofluorescent staining results, it may be important to maximize signal over background by using a relatively bright fluorochrome-antibody conjugate (Cat. No. 106306) or by using a high sensitivity, three-layer staining technique (e.g., including a biotinylated anti-Armenian hamster IgG (Cat. No. 405501) second step, followed by SAv-PE (Cat. No. 405204)). The Ultra LEAF? purified antibody (Endotoxin < 0.01 EU/?g, Azide-Free, 0.2 ?m filtered) is recommended for functional assays (Cat. No. 106327).

TotalSeq? reagents are designed to profile protein levels at a single cell level following an optimized protocol similar to the CITE-seq workflow. A compatible single cell device (e.g.?10x Genomics Chromium System and Reagents) and sequencer (e.g. Illumina analyzers) are required. Please contact?technical support?for more information, or visit?biolegend.com/totalseq.

The barcode flanking sequences are CGGAGATGTGTATAAGAGACAGNNNNNNNNNN (PCR handle), and NNNNNNNNNCCCATATAAGA*A*A (capture sequence). N represents either randomly selected A, C, G, or T, and * indicates a phosphorothioated bond, to prevent nuclease degradation.

View more applications data for this product in our?Scientific Poster Library.

1. Walunas TL, et al. 1994. Immunity 1:405. (Block, IP)
2. Cilio CM, et al. 1998. J. Exp. Med. 188:1239. (Block)
3. Issazadeh S, et al. 1999. J. Immunol. 162:761. (Block)
4. McCoy K, et al. 1997. J. Exp. Med. 186:183. (Block)
5. Hsu HC, et al. 2007. J. Immunol. 178:5357. (ELISA Capture)
6. Sugita S, et al. 2010. Invest. Ophthalmol. Vis. Sci. 51:5783. PubMed

Activated T cells and B cells

1. Barclay A, et al. 1997. The Leukocyte Antigen FactsBook Academic Press.
2. Allison JP, et al. 1995. Science 270:932.
3. Waterhouse P, et al. 1995. Science 270:985.
4. Linsley PS, et al. 1991. J. Exp. Med. 174:561.