pdf

Size: px

Start display at page:

Download "pdf"

せとかしもとり
6 years ago
Views:

1 BD FACSArray

2 BD FACSArray Unlimited Possibilities BD BD FACS BD FACSArray 96 2 BD FACSCalibur 4 Count BD Cytometric Bead Array CBA Flex Set ELISA Western Blot Propldi Ce BD Cytometric Bead Array CBA Flex Set NIR-A Red-A Multiplexed Bead Analysis

3 Red-A Count 0 50 Far Red-A Proliferation 10 6 um lodide Yellow-A (x 1,000) ll Cycle Analysis Propldium lodide Yellow-A Cell Viability Measurements Phospho-Histone A647 Red-A Propldium lodide Yellow-A (x 1,000) Cell Signaling CD19 PE Cy7 Far Red-A CD16+56 PE Yellow-A CD3 APC Red-A CD3 APC Red-A Cell Surface Staining BD FACSArray 4 Phenotype 96 2

4 BD Cytometric Bead Array (CBA) Flex Set BD CBA Flex Set BD FACS BD CBA Flex Set BD CBA BD CBA PE PE 1 PE BD CBA Flex Set 1 50 L 172 ELISA Western Blot Ready-to-Use BD 3

5 BD CBA Flex Set Human Mouse/Rat Cell Signaling BD CBA Flex Set FCAP Array Human Mouse/Rat Cell Signaling 3. 1 BD CBA Cell Signaling Flex Set 1 2 Cell Lysate 3 Assay Dilutions 4 BD CBA Cell Signaling Flex Set Capture 5 Capture50 L 6. FCAP Array 6 BD CBA Cell Signaling Flex Set PE 50 L 750 L L Wash Buffer 4

6 BD CBA Flex Set BD CBA Flex Set Western Blot 1 BD CBA Cell Singnaling Flex Set Western Blot Western Blot CV% Phospho-ZAP-70 c-jun 1 anti-cd3anti-cd28 anti-lg 2 Jurkat SDS 1% 1 Jurkat 2 2 units/ml2 2 anti-cd3anti-cd28 T Jurkatanti-CD3anti-CD g 9 BD CBA Flex Set ERK, JNK, p38, PLC, ZAP-70, Itk, LAT, c-jun, RSK units/ml SDS anti-phospho-zap-70 Western Blot BD CBA Flex Set Western Blot BD CBA Flex Set ZAP

median fluorescence intensity 100 MFI2 PD-98059 MEK ERK RSK PD-98059 ERK RSK 3 JurkatPP2 Lck

7 1 2 3 Control MEK Inhibitor (PD-98059) Src Family Kinase Inhibitor (PP2) 3 T Jurkat 200 M PD MEK PP2 Src family tyrosine kinase 20 anti-cd3anti-cd28 21 BD CBA Flex Set MFI median fluorescence intensity 100 MFI2 PD MEK ERK RSK PD ERK RSK 3 JurkatPP2 Lck T Western Blot 4 MAP 1 BD CBA Flex Set MAP p38, ERK, JNK Western blot BD CBA Flex Set Western Blot 6

8 BD CBA Flex Set FCAP Array TM BD FACSArray BD CBA Flex SetFCAP Array BD CBA Flex Set BD CBA Flex Set 72 Easy-to Use Fitting Accuracy 7

9 96 MFI Fitting Accuracy Microsoft Excel MFI SDMicrosoft Excel 21 CFR Part11 8

10 BD FACSArray 532nm635nm nm Yellow Far Red PE DsRed PI * PE-Cy7 PerCP-Cy5.5 PerCP PE-Cy5 ** 7-AAD 635nm Red NIR APC Alexa 647 APC-Cy7 PI Far Red * PE-Cy5 635nm 660nm APC ** 1) Annexin V PS PS Annexin V kda Ca 2 PS PS Annexin V Annexin V 7-Amino-Actinomycin D (7-AAD) Annexin V7-AAD Annexin V 7-AAD BD FACSArray SSC-A (x 1,000) 7-AAD Far Red-A 0 5 Sample_001_7-AAD/Annexin FSC-A (x 1,000) Sample_001_7-AAD/Annexin Annexin-VAPC Red-A 7-AAD Far Red-A 0 Sample_001_7-AAD/Annexin Annexin-VAPC Red-A 4 BD FACSArray PE PE-Cy7 APC APC-Cy7 4 CD45RA PE CD3 PE-Cy7 CD4 APC CD8a APC-Cy7 BD FACSArray P1: CD3 vs CD45RA CD3 CD45RA P2 CD3 vs CD4 Q2 CD3 CD4 CD3 vs CD8a Q2 CD3 CD8a CD8 vs CD45RA CD8 CD45RA CD8a APC-Cy7 NIR-A CD45RA PE Yellow-A 0 0 mouse spleen 4color-Well CD3 PE-Cy7 Far Red-A mouse spleen 4color-Well CD4 APC Red-A 0 CD45RA PE Yellow-A 0 mouse spleen 4color-Well CD3 PE-Cy7 Far Red-A mouse spleen 4color-Well 6 CD3 PE-Cy7 Far Red-A CD8a APC-Cy7 NIR-A 9

PIDNA 2) PI3 G0 /G1 S G2/M Anti-Histone H3 (pser28) - Alexa Fluor 647 G2 M PI Yellow P5 PIG2/M M SSC-A (x 1,000) Sample_001_Cell Cycle FSC-A (x 1,000) SSC-A (x 1,000) Sample_001_Cell Cycle 2 10 10 3

Sample_001_Well_001 SSC-A (x 1,000) Count 0 300 Sample_001_Well_001 50 100 150 7AAD Far Red-A (x 1,000) Sample_001_Well_001 0 10 BrdU APC Red-A 10 3 10 4 10 5 Red-A (x 1,000) Count 0 25 50 75 100 2 7

11 PIDNA 2) PI3 G0 /G1 S G2/M Anti-Histone H3 (pser28) - Alexa Fluor 647 G2 M PI Yellow P5 PIG2/M M SSC-A (x 1,000) Sample_001_Cell Cycle FSC-A (x 1,000) SSC-A (x 1,000) Sample_001_Cell Cycle Red-A Sample_001_Cell Cycle Sample_001_Cell Cycle DNA 3) DNA BrdU bromodeoxyuridine Thymidine BrdU anti- BrdU SDNA 8 BD APC BrdU Flow Kit DNA BrdU CD8 IL-2 CD8 CD8 FSC-A (x 1,000) Sample_001_Well_001 SSC-A (x 1,000) Count Sample_001_Well_ AAD Far Red-A (x 1,000) Sample_001_Well_ BrdU APC Red-A Red-A (x 1,000) Count Yellow-A (x 1,000) Yellow-A (x 1,000) Sample_001_Well_001 CD8 APC-Cy7 NIR-A 0 7AAD Far Red-A (x 1,000) 7AAD Far Red-A (x 1,000) Sample_001_Well_001 Sample_001_Well_001 CD8 APC-Cy7 NIR-A 0 CD8 APC-Cy7 NIR-A 0 BrdU APC Red-A IL-2 PE Yellow-A 8 1) BD FACSArray 2) BD FACSArray 3) BD FACSArray BrdU 10

12 BD FACSArray Easy-to Use Workspace Acquire Workspace Analyze Workspace Set Up Workspace Prepare Workspace BD FACSArray BD FACSArray 11

13 12

14 BD CBA 1. Bishop, J.E. and K.A. Davis A flow cytometric immunoassay for ß2-microglobulin in whole blood. J. Immunol. Methods 210: Camilla C., J.P. Defoort, M. Delaage, R. Auer, J. Quintana, T. Lary, R. Hamelik, S. Prato, B. Casano, M. Martin and V. Fert A new flow cytometry-based multi-assay system. 1. Application to cytokine immunoassays. Cytometry Suppl. 8: Carson, R., and D. Vignali Simultaneous quantitation of fifteen cytokines using a multiplexed flow cytometric assay. J. Immunol. Methods 227: Chen, R., L. Lowe, J.D. Wilson, E. Crowther, K. Tzeggai, J.E. Bishop and R. Varro Simultaneous quantification of six human cytokines in a single sample using microparticle based flow cytometric technology. Clin. Chem. 9: Kricka, LJ Simultaneous multianalyte immunoassays. In Immunoassay. Diamandis, E.P. and T.K. Christopoulos, eds. Academic Press. pp Lund-Johansen, F., K. Davis, J. Bishop and R. de W. Malefyt Flow cytometric analysis of immunoprecipitates: High-throughput analysis of protein phosphorylation and proteinprotein interactions. Cytometry 39: McHugh, T.M Flow microsphere immunoassay for the quantitative and simultaneous detection of multiple soluble analytes. Methods in Cell Biology 42: Oliver, K.G., J.R. Kettman and R.J. Fulton Multiplexed analysis of human cytokines by use of the FlowMetrix system. Clin. Chem. 44: Stall, A., Q. Sun, R. Varro, L. Lowe, E. Crowther, B. Abrams, J. Bishop, and K. Davis A single tube flow cytometric multibead assay for isotyping mouse monoclonal antibodies. Abstract LB77. Experimental Biology Meeting 1998 (late-breaking abstracts). 10. Cook, E.B., J.L. Stahl, L. Lowe, R. Chen, E. Morgan, J. Wilson, R. Varro, A. Chan, F.M. Graziano, and N.P. Barney Simultaneous measurement of six cytokines in a single sample of human tears using microparticle-based flow cytometry: allergics vs. non-allergics. J. Immunol. Methods. 254: Dotti, G., B. Salvoldo, S. Takahashi, T. Goltsova, M. Brown, D. Rill, C. Rooney, and M. Brenner Adenovector-induced expression of human-cd40-ligand (hcd40l) by multiple myeloma cells: A model for immunotherapy. Exp. Hematol. 29:

16 BD FACSArray BD FACSArray PE 200 MESF CV 7% Spheritech Rainbow Particle Cat L m 10 mw 532 nm 10 mw 635 nm m N/A m 430 m 523nm PMT1 532 nm PMT nm 685 nm 635 nm PMT nm nm L 20 L 3596 BD CBA L 2 L sec MultiScreen L 2 L sec BD Falcon cat % 4L L BD FACS Sheath Solution cm 64.3 cm 49.8 cm 39 kg 100/115/230 VAC 15 A 50/60Hz 500W 79 cm 61 cm 64 cm 51kg Microsoft Windows Microsoft Corporation Pentium Multiscreen Millipore Corporation BD, BD BD FACS BD FACSArray BD FACSCalibur BD Falcon BD R

CBA Kit Manual Human

CBA Kit Manual Human BD Pharmingen Cytometric Bead Array (CBA) Kit Manual Human Th1/Th2 Cytokine CBA BD Biosciences Section 1 1 Multiplexing BD Cytometric Bead Array CBA Multiplexing BD CBA CBA ELISA ELISA BD Human Th1/Th2