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Single cell a chip kit

Manufactured by 10x Genomics
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Sourced in United States

The Single Cell A Chip Kit is a lab equipment product from 10x Genomics. It is designed for single-cell analysis, providing a platform for processing and analyzing individual cells. The core function of this kit is to enable the isolation, encapsulation, and processing of single cells for downstream genomic analysis.

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Lab products found in correlation

Single cell 3 library gel bead kit v2, by 10x Genomics (2 mentions) Gemcode single cell instrument, by 10x Genomics (2 mentions) I7 multiplex kit, by 10x Genomics (2 mentions) Hiseq 2500, by Illumina (2 mentions) Dynabeads myone silane beads, by Thermo Fisher Scientific (2 mentions) S1000tm touch thermal cycler, by Bio-Rad (1 mentions) Chromium single cell 3 library and gel bead kit v2, by 10x Genomics (1 mentions) Chromium single cell controller, by 10x Genomics (1 mentions) Agilent 4200, by Agilent Technologies (1 mentions) Chromium single cell 5 library construction kit, by 10x Genomics (1 mentions) Chromium single cell 5 library gel bead kit v1, by 10x Genomics (1 mentions) Single cell 3 library and gel bead kit v2, by 10x Genomics (1 mentions) Chromium single cell 3 library, by 10x Genomics (1 mentions) Nextseq 500, by Illumina (1 mentions) Countess 2, by Thermo Fisher Scientific (1 mentions) Agilent hs dna chip, by Agilent Technologies (1 mentions) Spriselect, by Beckman Coulter (1 mentions) Novaseq platform, by Illumina (1 mentions) Single cell 3 library, by 10x Genomics (1 mentions)

8 protocols using single cell a chip kit

1

Single-cell RNA-seq Library Preparation

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The Chromium Single Cell 3′ Library and Gel Bead Kit V2 (10x Genomics, 120237) and Single Cell A Chip Kit (10x Genomics, 120236) were used for cell capture. The cell suspension (300–600 living cells per microlitre, as determined by Count Star) was loaded onto the Chromium Single Cell Controller (10x Genomics) to generate single-cell gel beads in emulsion (GEMs) according to the manufacturer’s protocol. In short, single cells were suspended in PBS containing 0.04% BSA. Approximately 3173 cells were added to each channel, and the target cell recovery was estimated to be ~16,544 cells in total. Captured cells were lysed, and the released RNA was barcoded through reverse transcription in individual GEMs. Reverse transcription was performed on a S1000TM Touch Thermal Cycler (Bio Rad) at 53 °C for 45 min, followed by 85 °C for 5 min and a hold at 4 °C. cDNA was generated and then amplified, and quality was assessed using an Agilent 4200 (performed by CapitalBio Technology, Beijing).
Wang X., Ning Y., Zhang P., Poulet B., Huang R., Gong Y., Hu M., Li C., Zhou R., Lammi M.J, & Guo X. (2021). Comparison of the major cell populations among osteoarthritis, Kashin–Beck disease and healthy chondrocytes by single-cell RNA-seq analysis. Cell Death & Disease, 12(6), 551.
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2

Single-Cell RNA-Seq Using 10x Genomics

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10x experiments were performed using the GemCode Single-Cell Instrument, Single Cell 3' Library & Gel Bead Kit v2 and Single Cell A Chip Kit (10x Genomics, Pleasanton, CA, USA) following the manufacturer’s protocol (CG00052_SingleCell3_ReagentKitv2UserGuide_RevD). In brief, the single cell suspension was resuspended in PBS and about 9,000 cells were loaded in one lane of the chip. Nanoliter-scale Gel bead-in-EMulsions (GEMs) were generated, mRNA reverse transcribed and cDNA amplified using 10 PCR cycles. The final library was PCR-amplified for 14 cycles and showed an average size of about 500 bp. Paired-end sequencing was carried out with the NextSeq 550 instruments at the DKFZ Genomics and Proteomics Core Facility (Heidelberg, Germany).
Bageritz J., Willnow P., Valentini E., Leible S., Boutros M, & Teleman A.A. (2019). Gene expression atlas of a developing tissue by single cell expression correlation analysis. Nature methods, 16(8), 750-756.
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3

Single-cell RNA-seq and TCR profiling protocol

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Cell suspensions (700–1200 living cells/μL determined by trypan blue staining) were loaded on a 10x Genomics ChromiumTM Single Cell Controller Instrument (10x Genomics, Pleasanton, CA, USA) to generate single-cell gel beads in emulsions (GEMs) by using Chromium Single Cell 5′ Library & Gel Bead Kit v1.0 (10x Genomics, Cat# 1000006) and Single Cell A Chip Kit (10x Genomics, Cat# 1000152). Captured cells were lysed and the released RNA was barcoded through reverse transcription in individual GEM. Barcoded cDNAs were pooled and cleanup by using DynaBeads MyOne Silane Beads (Thermo Fisher, Cat# 37002D), and then amplified and cleanup for further next-generation library construction. Single-cell RNA-seq libraries were prepared using Chromium Single Cell 5′ Library & Gel Bead Kit v1.0 (10x Genomics, Cat# 1000006) and i7 Multiplex Kit (10x Genomics, Cat# 120262), following the manufacturer’s instructions.
TCR libraries were prepared using Chromium Single Cell V(D)J Enrichment Kit, Human T Cell (10x Genomics, Cat# 1000005), Chromium Single Cell 5′ Library Construction Kit (10x Genomics, Cat# 1000020), and i7 Multiplex Kit (10x Genomics, Cat# 120262), following the manufacturer’s instructions. Sequencing was performed on an Illumina HiSeq X Ten (Illumina, San Diego, CA, USA) with pair end 150 bp (PE150).
Hui Z., Zhang J., Ren Y., Li X., Yan C., Yu W., Wang T., Xiao S., Chen Y., Zhang R., Wei F., You J, & Ren X. (2022). Single-cell profiling of immune cells after neoadjuvant pembrolizumab and chemotherapy in IIIA non-small cell lung cancer (NSCLC). Cell Death & Disease, 13(7), 607.
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4

Single-Cell RNA-Seq Profiling Protocol

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Each cell suspension was subjected to 3′ single-cell RNA sequencing using Single Cell A Chip Kit, Single Cell 3′ Library and Gel Bead Kit V2, and i7 Multiplex Kit (10x Genomics, Pleasanton, CA, USA) with a cell recovery target of 5000, following the manufacturer’s instructions. Libraries were sequenced on an Illumina HiSeq2500, and mapped to the GRCh38 human reference genome using the Cell Ranger toolkit (version 2.1.0).
Kim N., Kim H.K., Lee K., Hong Y., Cho J.H., Choi J.W., Lee J.I., Suh Y.L., Ku B.M., Eum H.H., Choi S., Choi Y.L., Joung J.G., Park W.Y., Jung H.A., Sun J.M., Lee S.H., Ahn J.S., Park K., Ahn M.J, & Lee H.O. (2020). Single-cell RNA sequencing demonstrates the molecular and cellular reprogramming of metastatic lung adenocarcinoma. Nature Communications, 11, 2285.
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5

Single-cell RNA-seq of Ulcerative Colitis

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Single cells were processed through the 10X Chromium Single-Cell Platform using the Chromium Single-Cell 3′ Library, Gel_Bead_Kit_v2 and the Single-Cell A Chip Kit (10X Genomics). Libraries were sequenced on Illumina NextSeq 500. We analyzed paired inflamed rectum (n = 4) and uninflamed sigmoid (n = 5) from 4 UC patients. Sequences were aligned to the GRCh38 using the Cell Ranger v.2.1.0 (10X). We excluded genes expressed in fewer than three cells, cells that expressed fewer than 500 genes, and unique molecular identifier (UMI) count less than 500 or greater than 60k. Per cell, we normalized by dividing UMI per gene by total UMI and log-transforming. We used Seurat (Seurat_R_package_v.3.0.1) integrated model to generate a combined UC model with cells from both inflamed and uninflamed samples retaining their group identity. We used shared nearest neighbor graph-based clustering with 15 principal components. UMAP visualizations, violin plots were produced using Seurat318 functions in conjunction with ggplot2. We conducted DEG in mast cell cluster (cluster 15) using FindMarkers function in Seurat, which is based on Wilcoxon test.
Chen E., Chuang L.S., Giri M., Villaverde N., Hsu N.Y., Sabic K., Joshowitz S., Gettler K., Nayar S., Chai Z., Alter I.L., Chasteau C.C., Korie U.M., Dzedzik S., Thin T.H., Jain A., Moscati A., Bongers G., Duerr R.H., Silverberg M.S., Brant S.R., Rioux J.D., Peter I., Schumm L.P., Haritunians T., McGovern D.P., Itan Y, & Cho J.H. (2021). Inflamed ulcerative colitis regions associated to MRGPRX2-mediated mast cell degranulation and cell activation modules, defining a new therapeutic target. Gastroenterology, 160(5), 1709-1724.
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6

Single-Cell RNA-Seq of Cell Suspensions

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Cell suspensions were submitted to the University of Wisconsin (UW) Biotechnology Center (UWBC) for processing. Collected cells were concentrated and cell viability was further validated using the Countess ™ II (Invitrogen). A total of 56,000 cells (7000 for each of 8 samples) were targeted using the 10X Genomics V(D)J Single-Cell RNA-Seq pipeline. Briefly, GEMs were prepared using the Single Cell A Chip Kit (10X Genomics). Full-length cDNA cleanup was performed using DynaBeads Myone Silane beads (Invitrogen). Full-length cDNA was then amplified by PCR (13 cycles) and post cDNA amplification cleanup was performed using SPRIselect (Beckman Coulter) and QC was performed using Agilent HS DNA chips. Feature barcode (FB) PCR amplification was done in 9 cycles. Sample indexing was then performed using the Chromium i7 Plate N, Set A indices (10X Genomics). Gene expression (GEX), FB, and V(D)J libraries were then prepared respectively. Libraries meeting all quality control criteria were then sequenced using the Illumina NovaSeq platform at the UW Gene Expression Center in collaboration with the UWBC DNA Sequencing Facility, Madison, Wisconsin. Libraries were sequenced at the following depth (reads/cell): GEX: 85,000; V(D)J: 11,000; FB: 6,400, with a total of 53,000 cells submitted for sequencing.
Chasman D.A., Welch Schwartz R., Vazquez J., Chavarria M., Jenkins E.T., Lopez G.E., Tyler C.T., Stanic A.K, & Ong I.M. (2023). Proteogenomic and V(D)J Analysis of Human Decidual T Cells Highlights Unique Transcriptional Programming and Clonal Distribution. Journal of immunology (Baltimore, Md. : 1950), 211(1).
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7

Single-Cell RNA-Seq Using 10x Genomics

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10x experiments were performed using the GemCode Single-Cell Instrument, Single Cell 3' Library & Gel Bead Kit v2 and Single Cell A Chip Kit (10x Genomics, Pleasanton, CA, USA) following the manufacturer’s protocol (CG00052_SingleCell3_ReagentKitv2UserGuide_RevD). In brief, the single cell suspension was resuspended in PBS and about 9,000 cells were loaded in one lane of the chip. Nanoliter-scale Gel bead-in-EMulsions (GEMs) were generated, mRNA reverse transcribed and cDNA amplified using 10 PCR cycles. The final library was PCR-amplified for 14 cycles and showed an average size of about 500 bp. Paired-end sequencing was carried out with the NextSeq 550 instruments at the DKFZ Genomics and Proteomics Core Facility (Heidelberg, Germany).
Bageritz J., Willnow P., Valentini E., Leible S., Boutros M, & Teleman A.A. (2019). Gene expression atlas of a developing tissue by single cell expression correlation analysis. Nature methods, 16(8), 750-756.
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8

Single-Cell RNA-Seq of Mouse Models

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The single-cell suspensions were subjected to 3′ single-cell RNA sequencing aiming for target recovery of 5,000 cells (C57BL/6J or Ldlr−/−) and 3,000 cells (Apoe−/−) using Single Cell A Chip Kit, Single Cell 3’ Library, and Gel Bead Kit V2, and i7 Multiplex Kit (10x Genomics). Solutions pertaining to 3′ chemistry and v2 of 10x Genomics were used to barcode individual cells. Libraries were sequenced on an Illumina HiSeq2500 and mapped to the mouse genome (build mm10) using the Cell Ranger toolkit (v3.0.2). Single-cell RNA sequencing parameters were summarized in Supplementary Table 2.
Lee S.H., Kim N., Kim M., Woo S.H., Han I., Park J., Kim K., Park K.S., Kim K., Shim D., Park S.E., Zhang J.Y., Go D.M., Kim D.Y., Yoon W.K., Lee S.P., Chung J., Kim K.W., Park J.H., Lee S.H., Lee S., Ann S.J., Lee S.H., Ahn H.S., Jeong S.C., Kim T.K., Oh G.T., Park W.Y., Lee H.O, & Choi J.H. (2022). Single-cell transcriptomics reveal cellular diversity of aortic valve and the immunomodulation by PPARγ during hyperlipidemia. Nature Communications, 13, 5461.
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