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400995 Immunomagnetic Beads

Immunomagnetic Beads: Ficha de datos de seguridad (MSDS o SDS), certificado de análisis y de calidad (CoA y CoQ), expedientes, folletos y otros documentos disponibles.
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      Descripción

      Replacement Information
      Description
      Overview

      This product has been discontinued.





      Each bead is approximately 1.3 µm in diameter with a magnetic core and thin polystyrene shell. Antibodies or proteins may be directly adsorbed or covalently attached via chemical modification of COOH surface groups. When coated with primary or secondary antibodies, these beads can be used in cell separation, cell sorting, immunoprecipitation and protein purification.
      Catalogue Number400995
      Brand Family Calbiochem®
      References
      Product Information
      FormBrown aqueous dispersion
      Formulation30 mg/ml solid content, supplied in distilled water.
      PreservativeNone
      Applications
      Application CommentsPreparation of Beads
      1. Thoroughly but gently mix the beads to achieve a uniform suspension. Transfer the desired amount of beads into another test tube. Use a 10% excess above the final desired quantity of beads to account for the amount of the fine beads that will be lost during the washing.
      2. Place the tube with the bead suspension on a test tube magnet. The magnet should capture the bulk of the beads immediately, while the smaller beads, or fines, tend to remain in suspension longer and will be the last to be captured. Pipette out the supernatant and the fines from the tube. Be careful during the pipetting not to bump or otherwise disturb the captured magnetic bead pellet. If you space the magnet so it does not touch the bottom of the container, you can capture the beads and pipette the supernatant and fines out of the tube without disturbing the captured pellet.
      3. Wash the beads to remove the fines from the bead preparation and equilibrate them in the coupling buffer. Add a volume of 50 mM borate buffer, pH 8.5, equal to the volume of the bead suspension transferred for washing. Remove the tube from the magnet; resuspend the beads in the borate buffer by vigorously vortexing the tube. Recapture the beads by placing the tube on a test tube magnet. Most of the beads will be captured, as before, and the fines will tend to remain in suspension. Pipette the supernatant and fines from the tube.
      4. Resuspend the beads in borate buffer, repeating the wash (step 3) 3-4 times. There will be fewer fines after every wash.
      5. Your yield should be 90% of the starting weight of beads and should be equivalent to your desired starting weight.

      Suggested Procedure for Adsorption of Antibodies to Immunomagnetic Beads

      Adsorption of Antibody onto the Beads
      1. Dialyze the antibody in PBS or phosphate buffer. Use 5 mg of beads/ml of coupling buffer-antibody solution. For affinity purified antibody, use 20-30 µg/mg (protein to beads). The antibody concentration of the coating solution will be 100 µg/ml. Most of the antibody (80-90%) will not bind to the beads during the coating step and can be recovered and used again.
      2. Add the appropriate amount of the antibody to the container first and mix the beads and antibody solution together to form a homogeneous suspension.
      3. Add half the final coupling volume of 100 mM borate buffer, pH 9.0. Again, mix the bead preparation to achieve homogeneous suspension.
      4. Bring the volume in the container to the final coupling volume by adding deionized water. Mix the final coating buffer suspension to achieve a homogeneous mixture.
      5. Affix the container to a rotator and rotate the suspension at room temperature for 16-18 h, adjusting the rotation speed so the protein does not foam (approximately 5 rpm).

      Washing After Adsorption of Antibody
      1. After the 16-18 h period for adsorption of the antibody to the solid phase, capture the beads and recover the supernatant. The supernatant will contain approximately 80% of the starting antibody that did not adsorb to the beads. You may wish to concentrate this antibody and keep it for subsequent use, particularly if the antibody is expensive or difficult to obtain.
      2. Resuspend the captured beads in 0.05% TWEEN® 20 detergent/PBS and vortex vigorously to suspend and wash the beads. Capture the beads and pipette the supernatant.
      3. Repeat the washing (step 2) three more times.

      Final Dilution and Blocking of the Beads
      1. After the beads have been washed, the captured beads will need to be resuspended in the final volume at a 3% (weight/volume) or 30 mg/ml of PBS, pH 7.4, with 0.1% BSA and 0.02% sodium azide. Substitute 0.01% merthiolate for sodium azide in bead preparations where the beads will be used to isolate living cells, such as in panning techniques. The final PBS/BSA/sodium azide solution should be passed through a 0.2 µm filter before adding to the beads.
      2. Add the PBS/BSA/sodium azide solution to achieve the final volume. Store at 4°C.

      Procedure for Covalent Coupling to Carboxylate Modified Immunomagnetic Beads

      Preparation of Protein
      1. Dialyze protein in 100 mM borate buffer, pH 9.0, overnight. Protein solution should be at a concentration of 2-5 mg/ml for optimum coupling.

      Covalent Coupling to the Beads
      1. Prepare coupling buffer (CDI buffer): 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide freshly prepared at 10 mg/ml in distilled water, pH 4.0. Repeat resuspension of beads in CDI buffer.
      2. Incubate for 2 h at room temperature or 10 min at 50°C.
      3. Capture the beads with the magnet and wash in 100 mM borate buffer, pH 9.0.
      4. Add protein solution to beads; incubate overnight at room temperature or for 10 minutes at 50°C.
      5. Recapture beads with magnet and remove supernatant. Only a small portion of the total protein in solution binds to the beads, so redialyze the supernatant into a neutral buffer.
      6. Resuspend the beads in PBS with 1% BSA or gelatin and 0.1% sodium azide. Store at 4°C.
      Biological Information
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Storage and Shipping Information
      Ship Code Ambient Temperature Only
      Toxicity Standard Handling
      Storage +2°C to +8°C
      Do not freeze Yes
      Special InstructionsStir once a week to prevent particle caking on the container bottom. Once settled, thoroughly resuspend the beads before using to restore separation and maximize reactive surface area.
      Packaging Information
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Número de referencia GTIN
      400995 0

      Documentation

      Immunomagnetic Beads Certificados de análisis

      CargoNúmero de lote
      400995
      Ficha técnica

      Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

      Revision06-August-2007 RFH
      DescriptionImmunomagnetic beads, 1.3 µm in diameter with a superparamagnetic core (12% magnetite) and a thin polystyrene shell, can be used for direct adsorption of proteins or for covalent attachment of ligands via -COOH surface groups. Use of these beads replaces or complements most other separation techniques. When coated with primary or secondary antibodies, applications include cell separation, cell sorting, DNA technology, immunoassay and protein purification. The beads respond to a magnetic field, demagnetizing when the field is removed. Because of their large surface areas and nonporous structure, they are separated from the liquid phase easily and rapidly.
      FormBrown aqueous dispersion
      Formulation30 mg/ml solid content, supplied in distilled water.
      PreservativeNone
      CommentsPreparation of Beads
      1. Thoroughly but gently mix the beads to achieve a uniform suspension. Transfer the desired amount of beads into another test tube. Use a 10% excess above the final desired quantity of beads to account for the amount of the fine beads that will be lost during the washing.
      2. Place the tube with the bead suspension on a test tube magnet. The magnet should capture the bulk of the beads immediately, while the smaller beads, or fines, tend to remain in suspension longer and will be the last to be captured. Pipette out the supernatant and the fines from the tube. Be careful during the pipetting not to bump or otherwise disturb the captured magnetic bead pellet. If you space the magnet so it does not touch the bottom of the container, you can capture the beads and pipette the supernatant and fines out of the tube without disturbing the captured pellet.
      3. Wash the beads to remove the fines from the bead preparation and equilibrate them in the coupling buffer. Add a volume of 50 mM borate buffer, pH 8.5, equal to the volume of the bead suspension transferred for washing. Remove the tube from the magnet; resuspend the beads in the borate buffer by vigorously vortexing the tube. Recapture the beads by placing the tube on a test tube magnet. Most of the beads will be captured, as before, and the fines will tend to remain in suspension. Pipette the supernatant and fines from the tube.
      4. Resuspend the beads in borate buffer, repeating the wash (step 3) 3-4 times. There will be fewer fines after every wash.
      5. Your yield should be 90% of the starting weight of beads and should be equivalent to your desired starting weight.

      Suggested Procedure for Adsorption of Antibodies to Immunomagnetic Beads

      Adsorption of Antibody onto the Beads
      1. Dialyze the antibody in PBS or phosphate buffer. Use 5 mg of beads/ml of coupling buffer-antibody solution. For affinity purified antibody, use 20-30 µg/mg (protein to beads). The antibody concentration of the coating solution will be 100 µg/ml. Most of the antibody (80-90%) will not bind to the beads during the coating step and can be recovered and used again.
      2. Add the appropriate amount of the antibody to the container first and mix the beads and antibody solution together to form a homogeneous suspension.
      3. Add half the final coupling volume of 100 mM borate buffer, pH 9.0. Again, mix the bead preparation to achieve homogeneous suspension.
      4. Bring the volume in the container to the final coupling volume by adding deionized water. Mix the final coating buffer suspension to achieve a homogeneous mixture.
      5. Affix the container to a rotator and rotate the suspension at room temperature for 16-18 h, adjusting the rotation speed so the protein does not foam (approximately 5 rpm).

      Washing After Adsorption of Antibody
      1. After the 16-18 h period for adsorption of the antibody to the solid phase, capture the beads and recover the supernatant. The supernatant will contain approximately 80% of the starting antibody that did not adsorb to the beads. You may wish to concentrate this antibody and keep it for subsequent use, particularly if the antibody is expensive or difficult to obtain.
      2. Resuspend the captured beads in 0.05% TWEEN® 20 detergent/PBS and vortex vigorously to suspend and wash the beads. Capture the beads and pipette the supernatant.
      3. Repeat the washing (step 2) three more times.

      Final Dilution and Blocking of the Beads
      1. After the beads have been washed, the captured beads will need to be resuspended in the final volume at a 3% (weight/volume) or 30 mg/ml of PBS, pH 7.4, with 0.1% BSA and 0.02% sodium azide. Substitute 0.01% merthiolate for sodium azide in bead preparations where the beads will be used to isolate living cells, such as in panning techniques. The final PBS/BSA/sodium azide solution should be passed through a 0.2 µm filter before adding to the beads.
      2. Add the PBS/BSA/sodium azide solution to achieve the final volume. Store at 4°C.

      Procedure for Covalent Coupling to Carboxylate Modified Immunomagnetic Beads

      Preparation of Protein
      1. Dialyze protein in 100 mM borate buffer, pH 9.0, overnight. Protein solution should be at a concentration of 2-5 mg/ml for optimum coupling.

      Covalent Coupling to the Beads
      1. Prepare coupling buffer (CDI buffer): 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide freshly prepared at 10 mg/ml in distilled water, pH 4.0. Repeat resuspension of beads in CDI buffer.
      2. Incubate for 2 h at room temperature or 10 min at 50°C.
      3. Capture the beads with the magnet and wash in 100 mM borate buffer, pH 9.0.
      4. Add protein solution to beads; incubate overnight at room temperature or for 10 minutes at 50°C.
      5. Recapture beads with magnet and remove supernatant. Only a small portion of the total protein in solution binds to the beads, so redialyze the supernatant into a neutral buffer.
      6. Resuspend the beads in PBS with 1% BSA or gelatin and 0.1% sodium azide. Store at 4°C.
      Storage +2°C to +8°C
      Do Not Freeze Yes
      Special InstructionsStir once a week to prevent particle caking on the container bottom. Once settled, thoroughly resuspend the beads before using to restore separation and maximize reactive surface area.
      Toxicity Standard Handling