rev: May 11, 2007

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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.


CD18 Antibodies


Mouse anti-human CD18 antibodies

-3 clones      CLB-LFA1/1,54 (purified & FITC)

                      Mem48 (purified, FITC & PE)

                      68-5A5 (purified, FITC & PE)            


CATALOG#                 CLONE#                  Workshop    Host         Form        Price

RDI-M1344clb              CLB-LFA1/1, 54            IV           mIgG1      purified     $375.00

RDI-M1669clb                 " "                                "                "             FITC        $375.00

RDI-CBL158                 MEM 48                        --            mIgG1       purified     $375.00

RDI-CBL158FT               " "                                --              "              FITC        $375.00

RDI-CBL158PE               " "                                --              "              PE            $469.00

RDI-CBL514                 68-5A5                           III           mIgG2a     purified      $375.00


Product Specification: mouse monoclonal anti-human CD18


PeliCluster  CD18

CAT#RDI- M1344clb

Test/vial 200

Clone CLB-LFA-1/1, 54

This clone has been derived from hybridization of SP2/0 cells with spleen cells of a (B10 x DBA/2)F1 mouse immunized with patient cells (3). This antibody has been clustered to CD18 in one of the international Workshop on Human White Cell differentiation Antigens

Isotype Mouse, IgG1.

Source Ascites fluid.

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 CD18- antigen (Lymphocyte Function associated Antigen-1), which is expressed on mature immunocompetent lymphocytes and their neoplastic counterparts, granulocytes and monocytes (2).

Molecular mass 95 kDa (1).

Application The monoclonal antibody can be used to detect LFA-1/MO1 deficiency in patients suffering from recurrent
bacterial infections.


Methods Indirect immunofluorescence staining with analysis by flowcytometry or fluorescence microscopy.


References

1. Miedema, F. et al., J. Immunol., 134, 3075 (1985).

2. Miedema, F. et al., Leukaemia Res., 9, 1099 (1985).

3. Miedema, F. et al., Eur. J. Immunol., 14, 518 (1984).


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.


FOR IN VITRO RESEARCH USE ONLY


Product Specification: mouse monoclonal CD18,conjugated to Fluorescein


PeliCluster  CD18 F


cat#RDI-M1669clb

Test/vial 100

Clone CLB-LFA-1/1, 54
This clone has been derived from hybridization of SP2/0 cells with spleen cells of a (B10 x DBA/2)F1 mouse immunized with patient cells (3). This antibody has been clustered to CD18 in one of the international Workshop on Human White Cell differentiation Antigens

Isotype Mouse, IgG1.


Source Ascites fluid.


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 CD18- antigen (Lymphocyte Function associated Antigen-1), which is expressed on mature immunocompetent lymphocytes and their neoplastic counterparts, granulocytes and monocytes (2).

Molecular mass 95 kDa (1).

Application The monoclonal antibody can be used to detect LFA-1/MO1 deficiency in patients suffering from recurrent bacterial infections.


Methods Direct immunofluorescence staining with analysis by flowcytometry or fluorescence microscopy.


References

1. Miedema, F. et al., J. Immunol., 134, 3075 (1985).

2. Miedema, F. et al., Leukaemia Res., 9, 1099 (1985).

3. Miedema, F. et al., Eur. J. Immunol., 14, 518 (1984).


APPLICATION  in direct Immunofluorescence 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

- Microtiter 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 microtiter 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 lymphocyte 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


Mouse ANTI-HUMAN CD18 ANTIGEN

PRODUCT CODE :RDI-CBL158   $375.00/vial 200ug

     Also available: FITC labeled   cat#RDI-CBL158FT    $438.00/vial

                               PE labeled    cat#RDI-CBL158PE   $469.00/vial


CLONE: MEM 48

ISOTYPE: IgG1

SOURCE: Mouse ascitic fluid

Immunizing Antigen: Peripheral blood leucocytes from an LGL-type leukaemia

Fusion Partner: Sp 2/0-AG 14 myeloma cell line

PURIFICATION METHOD: ion exchange chromatography

SPECIFICITY: This antibody recognizes the CD18 molecule, a 95 Kda integrin molecule (B2 integrin) that forms the common beta chain of the CD11/CD18 heterodimer integrins which are expressed on all leucocytes. the B2 integrin is the ICAM 1 receptor.

APPLICATIONS: * Identification of CD18+ leucocytes by direct immunofluorescence staining of viable cell suspensions.

* Identification of CD18+ leucocytes present within cryostat sections of frozen tissue.

* Studies of cell-cell adhesion. This antibody will block cell-cell interaction.

* Suitable for western blotting studies

REFERENCES: Bazil, V. et al. Fol. Biol. (Prague) 36 (1990)

                          Larson, R.S. & Springer T.A. Immunol Rev. 114, 181-217 (1990)

                          Stefanova I & Horejsi V. J. Immunol 5 1587-1592 (1991)


PRESENTATION: Each monoclonal is delivered in a sealed vial at a concentration of not less than 200µg/2ml in phosphate buffered saline containing 10mM sodium azide and 1mg/ml of bovine serum albumin. We recommend that each laboratory determine an optimum working titre for use in its particular application.

STORAGE: Purified monoclonal antibodies are generally stable for long periods at 4oC. However repeated warming to room temperature and re-cooling may result in a loss of activity and hence effective working titre. It is recommended that monoclonal antibodies are aliquotted upon receipt and then stored at -20C. After thawing they should then be used as a stock solution for up to 30 days. Dilute working solutions are best prepared on the day of use.


For research use only. Not supplied for use in human diagnostic or therapeutic procedures


Mouse Monoclonal Antibody To: Human CD18

Product CAT#:                          RDI-CBL514         $375.00

Also available: FITC labeled cat#RDI-CBL514FT   $375.00/100 test vial

                           PE labeled cat#RDI-CBL514PE   $469.00/100 test vial

Clone: 68-5A5

Isotype: IgG2a

Immunizing Antigen: Peripheral blood mononuclear cells.

Fusion Partner: NS1 myeloma cell line.

Source: Tissue culture supernatant

Purification Method: Protein A affinity chromatography

Specificity: This antibody recognizes the CD18 molecule, a 95 kDa integrin molecule (B2 integrin) that forms the common beta chain of the CD11/CD18 heterodimer integrins which are expressed on all leucocytes.

Applications: *Identification of CD18+ leucocytes present on cryostat sections

                     *Studies of cell-cell and cell-matrix adhesion.

                     *Suitable for western blotting studies.

                     *Flow Cytometry.

PRESENTATION: Each monoclonal is delivered in a sealed vial at a concentration of not less than 200µg/2ml in phosphate buffered saline containing 10mM sodium azide and 1mg/ml of bovine serum albumin. We recommend that each laboratory determine an optimum working titre for use in its particular application


STORAGE: Purified monoclonal antibodies are generally stable for long periods at 4oC. However repeated warming toroom temperature and re-cooling may result in a loss of activity and hence effective working titre. It is recommended that monoclonal antibodies are aliquotted upon receipt and then stored at -200C. After thawing they should then be used as a stock solution for up to 60 days. Dilute working solutions are best prepared on the day of use.


For research use only. Not supplied for use in human diagnostic or therapeutic procedures


Also available in BULK, without BSA and/or without azide

cat#RDI-CBL514-1X $1062.00/1mg or without azide= cat#RDI-CBL514-1XP


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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