rev: January 4, 2005
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CD CLUSTERED
ANTIBODIES
(anti-Human and others as indicated)
RDI Division of Fitzgerald Industries Intl offers a wide line of antibodies. Since no one antibody works best for all applications (flow cytometry, neutralization, blotting, histochemistry, ELISA, etc), we offer many different types of antibodies to help solve this problem. Please inquire for other applications or types of antibodies not listed below. All products are for in vitro research use only-not for use in or on humans or animals-not for use in diagnostics. Price/availability/specifications subject to change without notice.
CD9 Antibodies
Mouse anti-human CD9 antibodies
-4 clones CLB-Trom/8,4E1 (purified & FITC)
MM2/57 (purified, FITC & PE)
SN4/C3-3A2 (purified, or Biotin or FITC or PE)
72F6
CATALOG# CLONE# Workshop Host Form Price
RDI-M1362clb CLB-trom/8, 4E1 IV mIgG2a purified $375.00
RDI-M1666clb " " " " FITC $375.00
RDI-CBL162 MM2/57 IV mIgG2b purified $375.00
RDI-CBL162FT " " " " FITC $375.00
RDI-CBL162PE " " " " PE $469.00
RDI-CD9abm-A2 SN4/C3-3A2 -- mIgG1k purified $312.00
RDI-CD9abm-A2BT " -- " Biotin $375.00/100ug
RDI-CD9abm-A2FT " -- " FITC $375.00/120T
RDI-CD9abm-A2PE " -- " PE $531.00/120T
RDI-CD9abm-72 72F6 --- mIgG1 TCS $688.00/1ml (paraffin)
PeliCluster CD9
cat#RDI- M1362clb $375.00/vial
Test/vial 200
Clone CLB-thromb/8, 4E1
This clone has been derived from hybridization of SP2/0 cells with spleen
cells of a BALB/c mouse immunized with human B-cell precursors (cALL+ - ALL).
This antibody has been clustered to CD9 in the second international Workshop
on Human White Cell differentiation Antigens
Isotype Mouse, IgG2a.
Source Ascites fluid of tumour bearing BALB/c mice.
Purification Ammoniumsulphate precipitation and ion exchange
chromatography.
Packing Each vial contains 1 ml with approximately 0.2 mg/ml monoclonal antibody
and 10 mg BSA in 20 mM TRIS and 150 mM NaCl, pH 8.0.
Preservative Mertiolate (0.001%).
Storage and stability Monoclonal antibodies should be stored in the dark
at 2-8°C. The reagent is stable until the expiry date stated on the
vial label.
Major reactivity The monoclonal antibody is directed against the CD9- antigen
(BA-2-antigen), which is expressed on human B- cell precursors, human platelets
and human monocytes. The monoclonal antibody reacts with early B-cell precursors,
pre-pre-B-cells, pre-B-cells, platelets and a subpopulation of monocytes.
Molecular mass 24 kDa.
Application Characterization of ALL and in some cases AML.
Functional studies on platelets.
Methods Indirect immunofluorescence staining with analysis by flowcytometry or fluorescence microscopy.
References 1. Reinherz, E.L., Haynes, B.F., Nadler, L.M., Bernstein, I.D.,
Leukocyte Typing II, Springer Verlag, New York, 2 (1985).
APPLICATION FOR AN INDIRECT IMMUNOFLUORESCENCE TECHNIQUE
Reagents
- FITC-conjugated goat anti-mouse immunoglobulin antiserum (M1204).
Remove aggregates by centrifugation at 1000 x g for 10 minutes.
- Buffer: Phosphate Buffered Saline (PBS), containing 0.2% Bovine Serum Albumin (BSA).
- PFA: Para-Formaldehyde 1%, pH 7.2, containing 0.2% BSA.
- Microtiter plates (96 wells, V bottom) or tubes.
Procedure
1 Fixate the cells with PFA 1% and wash them with buffer.
2 Prepare a mononuclear cell suspension with a concentration of 1 x 10 7 cells/ml in buffer.
3 Add 45 µl of cell suspension to the microtiter wells or tubes.
4 Add 5 µl monoclonal antibody to the microtiter wells or tubes and mix gently.
5 Incubate for 30 minutes at 2-8 C.
6 Add 100 µl buffer to the microtiter wells or 2 ml buffer to the tubes and centrifuge at 500 x g for 5 minutes.
7 Aspirate the supernatant from the cell pellet and resuspend the cells.
8 Add 200 µl buffer to the microtiter wells or 2 ml buffer to the tubes and centrifuge at 500 x g for 5 minutes.
9 Aspirate the supernatant from the cell pellet and resuspend the cells.
10 Add 50 µl FITC-conjugated goat anti-mouse immunoglobulin serum, solid phase adsorbed with human immunoglobulins, diluted 1:80.
11 Incubate for 30 minutes at 2-8 C.
12 Repeat step 6-9.
13 Add 200 µl buffer.
14 The number of positive cells can be determined by flowcytometry analysis or by means of fluorescence microscopy.
NOTE: Care should be taken when drawing blood to avoid activation of platelets.
Platelets activation markers are weakly expressed on platelets prepared from directly fixed normal whole blood.
The expression is maximal on thrombin (1 U/ml) stimulated washed platelets.
FOR IN VITRO RESEARCH USE ONLY
PeliClusterCD9 F cat#RDI- M1666clb $375.00/vial
Test/vial 100
Clone CLB-thromb/8, 4E1
This clone has been derived from hybridization of SP2/0 cells with spleen
cells of a BALB/c mouse immunized with human B-cell precursors (cALL+ - ALL).
This antibody has been clustered to CD9 in the second international Workshop
on Human White Cell differentiation Antigens
Isotype Mouse, IgG2a.
Source Ascites fluid of tumour bearing BALB/c mice.
Purification Ammoniumsulphate precipitation and ion exchange chromatography.
Conjugation Conjugated with fluorescein iso thiocyanate isomer 1 (FITC).
Molecular F/P ratio between 5.5 - 9.5.
Packing Each vial contains 1 ml FITC conjugated monoclonal antibody and 10
mg BSA in PBS.
Preservative Mertiolate (0.001%).
Storage and stability Monoclonal antibodies should be stored in the dark
at 2-8°C. The reagent is stable until the expiry date stated on the
vial label.
Major reactivity The monoclonal antibody is directed against the CD9- antigen (BA-2-antigen), which is expressed on human B- cell precursors, human platelets and human monocytes. The monoclonal antibody reacts with early B-cell precursors, pre-pre-B-cells, pre-B-cells, platelets and a subpopulation of monocytes.
Molecular mass 24 kDa.
Application Characterization of ALL and in some cases AML.
Methods Direc immunofluorescence staining with analysis by flowcytometry
or fluorescence microscopy.
References 1. Reinherz, E.L., Haynes, B.F., Nadler, L.M., Bernstein, I.D.,
Leukocyte Typing II, Springer Verlag, New York, 2 (1985).
For research only! Not for human use! M1666\01 0808961318
FOR IN VITRO RESEARCH USE ONLY
PRODUCT CODE :RDI-CBL162 $375.00/vial 200ug purified
Also available FITC labeled cat#RDI-CBL162FT $375.00/100 Test vial
PE labeled cat#RDI-CBL162PE $469.00/100 Test vial
CLONE MM2/57
ISOTYPE IgG2b
IMMUNIZING ANTIGEN Human platelet glycoprotein
FUSION PARTNER NS1 myeloma cell line
SOURCE Tissue culture supernatant
PURIFICATION METHOD Protein A affinity chromatography
SPECIFICITY
This antibody reacts with a 25 kDa molecular weight band on gels of non-reduced platelet membranes and immunoprecipitates a similar band from I125 labelled platelets. The antigen is expressed on eosinophils, granulocytes, monocytes and numerous stromal cells. This CD9 antigen belongs to the TM4 superfamily of cell surface proteins with the N- and C-terminals cytoplasmically located. This antibody reacts immunohistochemically on frozen sections of platelets, monocytes and pre-B cells with differential staining on lymphoid and epithelial tissues.
APPLICATIONS
*Characterization of leukaemias and lymphomas
*Bone marrow purging in autologous bone marrow transplantation
*Immunoprecipitation studies of the CD9 antigen
*Platelet inhibition and aggregation studies and association with CD41/CD61
KNOWN SPECIES CROSS REACTIVITY
Positive with rabbit platelets and fibroblasts and mouse
REFERENCES
Leucocyte Typing IV, Oxford University Press (1989)
Leucocyte Typing V, Oxford University Press (1995)
Boucheix, C. et al. J. Biol. Chem. 266, 117-122 (1990)
Lanza, F. et al. J. Biol. Chem. 266, 10638-45 (1990)
LOT SPECIFIC DATA
PRESENTATION The monoclonal is presented at a concentration of 200µg/2ml
in phosphate buffered saline containing 10mM sodium azide and 1mg/ml bovine
serum albumin. We recommend that each laboratory determine an optimum working
titre for use in its particular application.
STORAGE For use within 1 month of purchase store at +4oC, for long term storage
aliquot antibody into small volumes and store at -20oC.
For anti-rabbit see http://www.researchd.com/rbcds/rbcdmono.htm
For anti-mouse see http://www.researchd.com/mscdabs/mscdabs.htm
For Research Use Only. Not supplied for use in dia
cat# RDI-CD9abm-72 $688.00/vial
Specificity: Human CD9 antigen
Clone: 72F6
Ig Class: IgG1
Antigen: prokaryotic recombinant protein corresponding to the major extracellular
loop of the CD9 molecule
Hybridoma partner: Mouse myeloma (p3-NS1-Ag4-1).
Preparation: Lyophilized tissue culture supernatant containing 15mM sodium
azide. Reconstitute with 1ml of sterile distilled water.
USE: Effective on paraffin wax embedded tissue using 1mM EDTA pH8.0 buffer
solution combined with the high temperature antigen unmasking technique frozen
sections:acetone fixation
Protocol: Immunohistochemistry:Typical working dilution 1:20-1:40 after high
temperature antigen unmasking technique using 1mM EDTA pH8.0. 60 Minutes
incubation at 25'C. Standard ABC technique.
-frozen sections:1:75-1:150 with ABC detection
Positive controls: fibroadenoma
Staining pattern: Membrane
Storage and stability: Store unopened lyophilized antibody at 4'C. Under
these conditions, there is no significant loss in product performance up
to the expiry date indicated on the vial label. The reconstituted antibody
is stable for at least two months when stored at 4'C. For long term storage,
aliquot in non frost freze freezer, Avoid repeated freeze thaw cycles. Prepare
fresh working dilutions daily.
Stricly for in vitro research use only-Not for use in or on humans or animals-not
for use in diagnostics
CD9 antigen is a 24 to 27kda glycoprotein expressed on the surface of developing B lymphocytes, platelets, monocytes, eosinophils, basophils,s timulated T lymphocytes and by neurons and glial cells in the peripheral nervous system. It belongs to a family of membrane proteins termed tetraspanins which transverse the membrane four times. In pre-B cells and platelets, Cd9 antigen regulates cell activation and aggregation possibly through an association with the integrin CD41/Cd61 (GPIIb/GPIIIa). It also regulates cell motility in a variety of cell lines, and appears to be an important regulator of Schwann cell behaviour in peripheral nerve. In melanoma and breast cancer, CD9 antigen expression may indicate a favorable prognosis as expression has been shown to occur predominantly on primary, non-metastatic tumours.
Sample High Temperature Unmasking Technqiue for Paraffin Sections: Buffer=1mM EDTA (ph 8.0)
1) Cut and mount sections on slides coated with "apes"
2) Deparrafinize sections and rehydrate to distilled water
3) Bring 1600ml 1mM EDTA* (pH 8.0) to a boil in a Prestige/Presto stainless
steel pressure cooker, using a hot plate. Cover but do not lock lid.
4) Position slides into metal staining racks and lower into pressure cooker
ensuring slides are well immersed in buffer Lock the lid. The small valve
will rise.
5) When the pressure indicator valve (the large one) has risen after about
4 minutes, Incubate sections for 1 minute.
6) After timer rings, Remove pressure cooker from heat source and run under
cold water with lid on. When the small valve sinks open lid and remove slides
and place immediately into distilled water. DO NOT OPEN LID UNTIL THE
SMALL VALVE Sinks or Vent Cover Lock Drops (see manual with your particular
unit).
7) Wash sections PBS buffer (10mM phosphate, 0.15M NaCl pH 7.5) for 2 X 5
minutes (or 100mM TRIS, 0.15M NaCl, pH 7.5)
8) Place sections in 1.5% hydrogen peroxide/methanol for 10 minutes (or use
other appropriate peroxidase blocking procedure).
9) Wash sections in distilled water for 2 X 5 minutes, then wash sections
in PBS buffer for 2 X 5 minutes.
10) Place sections in normal serum for 20 minutes.
11) Cover sections with primary antibody for at least 60 minutes at room
temp. (Optimum dilution, incubation time and temperature must be determined
for each antibody and in each lab).
12) Wash in PBS buffer for 2 X 5 minutes
13) Incubate sections in secondary antibody for 30 minutes
14) Wash in PBS buffer for 2 X 5 minutes
15) Incubate slides in ABC complex for 30 minutes
16) Wash in PBS buffer for 2 X 5 minutes
17) Incubate slides in DAB or other suitable peroxidase substrate
18) Wash in distilled water for 2 X 5 minutes
19) Counterstain with haematoxylin (if required), dehydrate, coverslip, and
mount
Material:
APES (3-aminopropyltriethoxysilane) cat#A3648 Sigma
To avoid sections becoming detached, sections should be mounted on "APES" covered slides, then dried at 37 DEG C overnight followed by drying at 56 DEG C for 60 minutes.
1mM EDTA (ph 8.0): Add 0.37g of EDTA (Sigma cat#E-5134) to 1 liter of distilled water. Adjust pH to 8.0 using 1.0M sodium hydroxide.
Tris: 100mmM Tris, 0.15M NaCl, pH 7.5.
PBS: 10mM Phosphate, 0.15M MaCl pH 7.5
RDI Division of Fitzgerald Industries Intl
34 Junction Square Drive
Concord MA 01742-3049
USA
phone (978) 371-6446 or (800) 370-2222
fax (978) 371-2266
EMAIL:antibodies@fitzgerald-fii.com
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