rev: 5/16/2008
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CD CLUSTERED ANTIBODIES
(anti-Human and others as indicated)
NOTE: 10% Discount on purchases of 3 or more any combination of CD antibodies. Please inquire for bulk or custom formulations (without preservative or carrier).
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.
CD66b
Mouse anti-human CD66b antibodies
-2 clone s
CATALOG# CLONE# Workshop Host Form Price
RDI-M1546clb CLB-B13.9
IV
mIgG1 purified
$412.50
new cat#: 10R-CD66baHU
RDI-M1594clb " " " FITC $375.00
RDI-CD66b-G10F5 G10F5 V mIgM purified $375.00
see sample protocols for conjugated antibody
Product Specification: mouse monoclonal anti-human CD66b
PeliCluster CD66b
cat#RDI- M1546clb
Test/vial 200
Clone CLB-B13.9
This clone has been derived from hybridization of SP2/0 cells with spleen
cells of a (BALB/c x A/J) mouse immunized with human granulocytes. This
antibody has been clustered to CD66b in the fifth international Workshop on
Human White Cell differentiation Antigens in
Isotype Mouse IgG1.
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 CD66b-
antigen, which is only expressed on the membrane of mature human neutrophil, eosinophil
and basophil granulocytes. After granulocyte activation the expression is
increased. The monoclonal antibody reacts with 100% of mature human
granulocytes. The monoclonal antibody does not react with normal human
peripheral B-cells, T-cells, monocytes and platelets.Molecular mass 100 kDa
(PI-linked protein).
Application Monitoring of mature granulocytes in peripheral blood.
Analysis of peripheral blood granulocytes from patients with Paroxysmal Nocturnal Haemoglobinuria (PNH).
Methods Indirect immunofluorescence staining with analysis by flowcytometry or fluorescence microscopy.
References 1. Schoot, C.E. van der, et al., Knapps, W. et al. (editors),
Leucocyte Typing IV, 840 (1989).
FOR IN VITRO RESEARCH USE ONLY
Product Specification: mouse monoclonal CD66b,conjugated to Fluorescein
PeliCluster CD66b F
cat#RDI-M1594clb
Test/vial 100
Clone CLB-B13.9 This clone has been derived from hybridization of SP2/0
cells with spleen cells of a (BALB/c x A/J) mouse immunized with human granulocytes.
This antibody has been clustered to CD66b in the fifth international Workshop
on Human White Cell differentiation Antigens in
Isotype Mouse IgG1.
Source Ascites fluid of tumour bearing BALB/c mice.
Purification Ammoniumsulphate precipitation and ion exchange chromatography.
Conjugation Cojugated 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 CD66b-
antigen, which is only expressed on the membrane of mature human neutrophil,
eosinophil and basophil granulocytes. After granulocyte activation the
expression is increased. The monoclonal antibody reacts with 100% of mature
human granulocytes. The monoclonal antibody does not react with normal human
peripheral B-cells, T-cells, monocytes and platelets.Molecular mass 100 kDa
(PI-linked protein).
Application Monitoring of mature granulocytes in peripheral blood.
Analysis of peripheral blood granulocytes from patients with Paroxysmal Nocturnal Haemoglobinuria (PNH).
Methods Direct immunofluorescence staining with analysis by flowcytometry or
fluorescence microscopy.
References 1. Schoot, C.E. van der, et al., Knapps, W. et al. (editors),
Leucocyte Typing IV, 840 (1989).
For research only!
APPLICATION in directImmunofluorescence techniques
Method with ficoll purified cells
Reagents and materials
- FITC-conjugated monoclonal antibodies.
- Buffer: Phosphate Buffered Saline (PBS), containing 0.2% Bovine Serum Albumine (BSA)
- PFA: Para-Formaldehyde 1%, pH 7.2, containing 0.2% BSA
- Microwell plates (96 wells, V bottom) or tybes.
Procedure
1 Prepare a mononuclear cell suspension with a concentratation of 1 x 10 7 cells/ml.
2 Ad 40 µl of cell suspension to microtiter wells or tubes.
3 Add 10 µl of the antibody to the microtiter wells or tubes and mix gently.
4 Incubate for 30 minutes at 2-8°C.
5 Add 150 µl buffer to the microtiter wells or 2 ml buffer to the tubes and centrifuge at 500 x g for 5 minutes.
6 Aspirate the supernatant from the cell pellet and resuspend the cells.
7 Add 200 µl buffer to the microtiter wells or 2 ml buffer to the tubes and centrifuge at 500 x g for 5 minutes.
8 Aspirate the supernatant from the cell pellet and resuspend the cells.
9 Cytoflow fluorometer analysis:
Add 200 µl buffer to the microplate wells and transfer this final cell suspension to appropriate testtubes, or add 200 µl buffer to the tubes.
10 If analysis within 8 hours is not possible add at no. 9, instead of buffer, 200 µl PFA 1%.
Fig. 1
Fluorescense profile of ficoll purified normal cells
(Scatter gates set on the granulocyte fraction)
WHOLE BLOOD METHOD
Due to sophisticated laser flowcytometry we are now able to distinguish
populations of human lymphocytes, monocytes and granulocytes by their
characteristics in different light scattering. By gating on the appropriate
cell cluster (Fig.2) we can easily determine mononuclear cells, which have been
immunofluorescently stained with monoclonal antibodies, and enumerate them.
Reagents and materials
- FITC-conjugated monoclonal antibodies.
- Lysing solution (NH 4Cl, pH 7.2).
- Buffer: PBS, containing 0.2 % BSA.
- PFA: Para-Formaldehyde 1%, pH 7.2, containing 0.2 % BSA.
- Tubes.
Procedure
1 Draw blood into a blood collection tube containing EDTA.
2 Deliver 100 µl (*) of well mixed whole blood to the bottom of the testtube.
3 Add 10 µl of the undiluted antibody to the bottom of the testtube, and mix gently.
4 Incubate for 30 minutes at 2-8°C or in an ice-water bath.
5 Mix the tubes and add 2 ml of lysing solution.
6 Incubate for 3-5 minutes at room temperature.
7 When lysing is completed, centrifuge the tubes at 500 x g for 5 minutes.
8 Aspirate the supernatant from the cell pellet, resuspend the cells in 1 ml buffer.
Place the tubes in the ice-water bath.
9 Analyse the samples within 8 hours.
If analysis within 8 hours is not possible centrifuge the tubes at 500 x g. Aspirate the supernatant from the cell pellet and resuspend the cells in 1 ml PFA 1%.
Fig. 2:
Scatter patern of a whole normal blood sample
R2 - lymphocytes R3 - monocytes R4 - granulocytes
Note:Use our special negative control to determine background fluorescence
produced due to Fc binding capacities by mononuclear cells.The concentration
and F/P ratio of our control has been adjusted to our FITC-conjugated
monoclonal antibodies.
* This method was developed for blood samples with a normal white count. It may
be necessary to adjust the quantity of blood for samples with very high or low
white counts
FOR IN VITRO RESEARCH USE ONLY
DATA SHEET: mouse anti-human CD66B clone G10F5
Catalog#: RDI-CD66B-G10F5 $281.00/vial
Package Size: 100ug purified in 02.ml (0.5mg/ml) 10mM sodium phosphate pH7.2,
500mM NaCl and 0.09% sodium azide. (sodium azide is toxic,dilute azide
compounds with copious amounts of water when disposing in drain to avoid
accumulation of explosive deposits-avoid contact with acids.)
Prep: purified from tissue culture supernatant via Protein G affinity
chromatography.
Species: mouse IgMk
CLONE: G10F5
Workshop Code: Workshop V, MA020
Activity: Reacts with a 100Kd, a glycosylphosphatidylinositol (GPI) anchored
protein expressed on granulocytes. This molecule was previously clustered as
CD67 in the IVth Workshop amd renamed CD66B in the Vth HLDA Workshop. CD66b is
a member of the CEA like glycoprotein family present on granulocytes and
referred to as non specific cross reacting antigenss. Granulocyte activation
induced with soluble stimulators (calcium ionophore, phorbol myristate acetate,
N- forylmethionyl-leucyl-phenylalanine)results in release and increased
expression of NCA. Findings suggest that these molecules may play a role in
phagocytosis, chemotaxis and adherence.
Application: -Indirect immunofluorescence cell surface staining ,
Immunohistochemistry on formalin-fixed paraffin embedded tisue sections using
citrate pretreatment and ABC/Dab detection. Recommend using a specific anti-IgM
secondary antibody.Titer for each application
References: 1) Schlossman, S. et al, ed.s 1995, Leucocyte Typing V, White Cell
Differentiation Antigens, Oxford University Press, NY
2) Eur J. Immunol. 23:2782 (1993)
3) J. Immunol. 151:4934 (1993)
4)
Journal of Leukocyte Biology 52:551.(1992)
Storage: Store at 4 DEG C.
Precautions:For In vitro research Use Only. Not for use in or on humans or
animals or or diagnostics.
Fitc labeled cat#RDI-CD66B-G10-FT $469.00/100 T vial
It is the responsibility of the user to comply with all local/state and Federal rules in the use of this product. We are not responsible for any patent infringements that might result with the use of or derivation of this product.
Precautions: For In vitro research Use Only. Not for use in or on humans or
animals or for diagnostics.
RDI Division of Fitzgerald Industries Intl
phone (978) 371-6446 or (800) 370-2222
fax (978) 371-2266
EMAIL:antibodies@fitzgerald-fii.com
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