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with, antigen, human, activation, CD69, CD19, from, cells, CD45, Typing, York,, present, eds., Dörken, blood, mouse, Testi, Leucocyte, White, Differentiation, Antigens., Oxford, University, Press;, receptor, that, expression, antibody, resting, Immunol.

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Becton, Dickinson and Company
BD Biosciences
2350 Qume Drive
San Jose, CA 95131 USA
01/2015 23-3109-04
bdbiosciences.com
ResearchApplications@bd.com
For Research Use Only. Not for use in diagnostic or therapeutic procedures.












Monoclonal
Antibodies
Detecting
Human
Antigens
BD FastImmune™
CD19/CD69/CD45
Catalog No. 340418 50 Tests 20 µL/test
RESEARCH
APPLICATIONS
Research applications include studies of:
• Activated B lymphocytes in peripheral blood1
• B-lymphocyte activation mechanisms2
DESCRIPTION
Specificity The CD19 (SJ25C1) antibody recognizes a 90-kilodalton (kDa) antigen that is present
on human B lymphocytes.3,4
The CD69 antibody recognizes a very early human lymphocyte activation antigen. The
CD69 antigen is a surface homodimer formed by the association of 28-kDa and 32-kDa
chains that are held together by disulfide bridges.1
The CD45 antibody recognizes a 180- to 220-kDa human leucocyte antigen that is a
member of the leucocyte common antigen (LCA) family.5
Antigen distribution The CD19 antigen is present on approximately 7% to 23% of human peripheral blood
lymphocytes (PBLs)6 and on splenocytes.7 CD19 is reactive with the B-lymphocyte areas
of normal tonsil and lymph nodes.8 The CD19 antigen is present on human
B lymphocytes at all stages of maturation but is lost on terminally differentiated plasma
cells.8 CD19 does not react with resting or activated T lymphocytes, granulocytes, or
monocytes.9
The CD69 antigen is present on activated T, B, and natural killer (NK) lymphocytes10
and platelets.11 CD69 is not expressed by resting PBLs.1 Upon activation, CD69 antigen
expression increases on lymphocytes; peak expression generally occurs within 18 hours,
preceding the appearance of HLA-DR, interleukin-2 (IL-2) receptor (CD25 antigen),
and transferrin receptor (CD71 antigen).12-14 CD69 and phorbol ester are comitogenic
for T lymphocytes.13 In thymus, the CD69 antigen is constitutively expressed on the
bright CD3+ subset.15
The CD45 antigen is present on all human leucocytes, including lymphocytes,
monocytes, granulocytes, eosinophils, and basophils in peripheral blood and has a role
in signal transduction, modifying signals from other surface molecules.5 CD45 reacts
weakly with mature circulating erythrocytes and platelets.5,16
Clones The CD19 antibody, clone SJ25C1, is derived from hybridization of SP2/0 mouse cells
with spleen cells from BALB/c mice immunized with NALM1 + NALM16 cells.
The CD69 antibody, clone L78, is derived from hybridization of Sp2/0-Ag14 mouse
myeloma cells with lymph node cells from BALB/c mice immunized with a CD8+
alloantigen-directed cytotoxic T-lymphocyte (CTL) cell line.17
23-3109-04 Page 2
The CD45 antibody, clone 2D1,18 is derived from hybridization of NS-1 mouse
myeloma cells with spleen cells from BALB/c mice immunized with human peripheral
blood mononuclear cells (PBMCs).
Composition The CD19, CD69, and CD45 antibodies are each composed of mouse IgG1 heavy
chains and kappa light chains.
The BD FastImmune™ reagent is supplied as a combination of CD19 FITC, CD69 PE,
and CD45 PerCP in 1.0 mL of phosphate-buffered saline (PBS) containing bovine
serum albumin (BSA) and 0.1% sodium azide.
PROCEDURE Visit our website (bdbiosciences.com) or contact your local BD representative for the
BD procedure Flow Cytometric Procedure for Assessing Lymphocyte Activation for the
complete activation protocol. This procedure follows the basic format described
previously. 19
1. After the activation process, pipet 20 µL of BD FastImmune CD19/CD69/CD45
into a labeled tube.
2. Add 50 µL of whole blood.
3. Vortex gently to mix and incubate for 15 minutes in the dark at room temperature.
4. Add 450 µL of 1X BD FACS™ lysing solution (Cat. No. 349202) to the tube.
5. Vortex gently and incubate for 15 to 30 minutes in the dark at room temperature.
REPRESENTATIVE DATA Flow cytometric analysis was performed on lysed whole blood with a gate (R1) set on
the CD45+ lymphocyte fraction. Laser excitation was at 488 nm.
Figure 1 Four-hour pokeweed mitogen (PWM)–activated CD45+ lysed whole blood analyzed with
a BD FACScan™ flow cytometer
HANDLING AND
STORAGE
Store vials at 2°C–8°C. Conjugated forms should not be frozen. Protect from exposure
to light. Each reagent is stable until the expiration date shown on the bottle label when
stored as directed.
WARNING All biological specimens and materials coming in contact with them are considered
biohazards. Handle as if capable of transmitting infection20,21 and dispose of with
proper precautions in accordance with federal, state, and local regulations. Never
pipette by mouth. Wear suitable protective clothing, eyewear, and gloves.
CHARACTERIZATION To ensure consistently high-quality reagents, each lot of antibody is tested for
conformance with characteristics of a standard reagent. Representative flow cytometric
data is included in this data sheet.
gating resting activated
CD45 PerCP CD19 FITC CD19 FITC
SS
C
C
D
69
P
E
C
D
69
P
E
R1
Page 3 23-3109-04
WARRANTY Unless otherwise indicated in any applicable BD general conditions of sale for non-US
customers, the following warranty applies to the purchase of these products.
THE PRODUCTS SOLD HEREUNDER ARE WARRANTED ONLY TO CONFORM TO THE QUANTITY AND CONTENTS
STATED ON THE LABEL OR IN THE PRODUCT LABELING AT THE TIME OF DELIVERY TO THE CUSTOMER. BD
DISCLAIMS HEREBY ALL OTHER WARRANTIES, EXPRESSED OR IMPLIED, INCLUDING WARRANTIES OF
MERCHANTABILITY AND FITNESS FOR ANY PARTICULAR PURPOSE AND NONINFRINGEMENT. BD’S SOLE LIABILITY
IS LIMITED TO EITHER REPLACEMENT OF THE PRODUCTS OR REFUND OF THE PURCHASE PRICE. BD IS NOT LIABLE
FOR PROPERTY DAMAGE OR ANY INCIDENTAL OR CONSEQUENTIAL DAMAGES, INCLUDING PERSONAL INJURY,
OR ECONOMIC LOSS, CAUSED BY THE PRODUCT.
REFERENCES 1. Testi R, D'Ambrosio D, De Maria R, Santoni A. The CD69 receptor: a multipurpose cell-surface trigger
for hematopoietic cells. Immunol Today. 1994;15:479-483.
2. Rasmussen A-M, Blomhoff H, Stokke T, Horejsi V, Smeland E. Cross-linking of CD53 promotes
activation of resting human B lymphocytes. J Immunol. 1994;153:4997.
3. Nadler LM. B Cell/Leukemia Panel Workshop: summary and comments. In: Reinherz EL, Haynes BF,
Nadler LM, Bernstein ID, eds. Leukocyte Typing II: Human B Lymphocytes. New York, NY: SpringerVerlag; 1986;2:3-43.
4. Moldenhauer G, Dörken B, Schwartz R, Pezzutto A, Knops J, Hammerling GJ. Analysis of ten B
lymphocyte-specific workshop monoclonal antibodies. In: Reinherz EL, Haynes BF, Nadler LM,
Bernstein ID, eds. Leukocyte Typing II: Human B Lymphocytes. New York, NY: Springer-Verlag;
1986;2:61-67.
5. Schwinzer R. Cluster report: CD45/CD45R. In: Knapp W, Dörken B, Gilks WR, et al, eds. Leucocyte
Typing IV: White Cell Differentiation Antigens. New York, NY: Oxford University Press; 1989:628-634.
6. Reichert T, DeBruyère M, Deneys V, et al. Lymphocyte subset reference ranges in adult Caucasians. Clin
Immunol Immunopath. 1991;60:190-208.
7. Tedder T, Zhou L-J, Engel P. The CD19/CD21 signal transduction complex of B lymphocytes. Immunol
Today. 1994;15:437-442.
8. Dörken B, Möller P, Pezzutto A, Schwartz-Albiez R, Moldenhauer G. B-cell antigens: CD19. In: Knapp
W, Dörken B, Gilks WR, et al, eds. Leucocyte Typing IV: White Cell Differentiation Antigens. New York,
NY: Oxford University Press; 1989:34-36.
9. Loken MR, Shah VO, Dattilio KL, Civin CI. Flow cytometric analysis of human bone marrow. II.
Normal B-lymphocyte development. Blood. 1987;70:1316-1324.
10. Schwarting R, Biedobitek G, Stein H. Cluster report: CD69. In: Knapp W, Dörken B, Gilks WR, et al,
eds. Leucocyte Typing IV: White Cell Differentiation Antigens. New York, NY: Oxford University Press;
1989:428-432.
11. Testi R, Pulcinelli F, Frati L, Gazzaniga P, Santoni A. CD69 is expressed on platelets and mediates platelet
activation and aggregation. J Exp Med. 1990;172:701-707.
12. Testi R, Philips J, Lanier LL. Constitutive expression of a phosphorylated activation (Leu-23) by CD3
bright thymocytes. J Immunol. 1988;141:2257.
13. Testi R, Philips JH, Lanier LL. Leu-23 induction as an early marker for functional CD3/T cell antigen
receptor triggering: requirement for receptor cross-linking, prolonged elevation of intracellular (Ca++),
and stimulation of protein kinase C. J Immunol. 1989;142:1854.
14. Chen JH, Prince H, Buck D, et al. Leu-23: an early activation antigen on human lymphocytes. Fed Proc.
1988;2:A1214.
15. Testi R, Philips J, Lanier LL. T cell activation via Leu-23 (CD69). J Immunol. 1989;143:1123-1128.
16. Ledbetter JA, Evans RL, Lipinski M, Cunningham-Rundles C, Good RA, Herzenberg LA. Evolutionary
conservation of surface molecules that distinguish T-lymphocyte helper/inducer and T
cytotoxic/suppressor subpopulations in mouse and man. J Exp Med. 1981;153:310-323.
17. Lanier LL, Buck DW, Rhodes L, et al. Interleukin 2 activation of natural killer cells rapidly induces the
expression and phosphorylation of the Leu-23 activation antigen. J Exp Med. 1988;167:1572.
18. Cobbold SP, Hale G, Waldmann H. Non-lineage, LFA-1 family, and leucocyte common antigens: new
and previously defined clusters. In: McMichael AJ, ed. Leucocyte Typing III: White Cell Differentiation
Antigens. New York, NY: Oxford University Press; 1987:788-803.
23-3109-04 Page 4
19. Maino VC, Suni MA, Ruitenberg JJ. Rapid Flow Cytometric Method for Measuring Lymphocyte Subset
Activation. Cytometry. 1994;20:127-133.
20. Protection of Laboratory Workers from Occupationally Acquired Infections; Approved Guideline —
Third Edition. Wayne, PA: Clinical and Laboratory Standards Institute; 2005. CLSI document M29-A3.
21. Centers for Disease Control. Perspectives in disease prevention and health promotion update: universal
precautions for prevention of transmission of human immunodeficiency virus, hepatitis B virus, and
other bloodborne pathogens in health-care settings. MMWR. 1988;37:377-388.
PATENTS AND
TRADEMARKS
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Company. © 2015 BD

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