I am getting high background using Dynabeads M-280 Streptavidin magnetic beads. How can I prevent this from happening?
The background might be caused by nonspecific binding to the BSA on the bead surface. Alternatively, high background might be caused by nonspecific binding to streptavidin. Increasing either the pH or the salt concentration might help reduce the binding. Dynabeads M-270 Streptavidin magnetic beads might be a better alternative; these beads are not coated with BSA and are hydrophilic, as they are based upon carboxylic acid chemistry.
What is the binding capacity of streptavidin-coupled Dynabeads magnetic beads?
- One milligram of Dynabeads M-280 Streptavidin magnetic beads typically binds 650-900 pmol of free biotin, 200 pmol of biotinylated peptide, up to 10 µg of biotinylated antibody, 10 µg of biotinylated double-stranded DNA, or 200 pmol of biotinylated single-stranded oligonucleotides.
- One milligram of Dynabeads M-270 Streptavidin magnetic beads typically binds more than 950 pmol of free biotin, 200 pmol of biotinylated peptide, up to 10 µg of biotinylated antibody, 10 µg of biotinylated double- stranded DNA, or 200 pmol of biotinylated single-stranded oligonucleotides.
- One milligram of Dynabeads MyOne Streptavidin C1 magnetic beads typically binds more than 2,500 pmol free biotin, 400 pmol of biotinylated peptides, up to 20 µg of biotinylated antibody, 20 µg of biotinylated double-stranded DNA, or 500 pmol of biotinylated single-stranded oligonucleotides.
-One milligram of Dynabeads MyOne Streptavidin T1 magnetic beads typically binds 1,100-1,700 pmol free biotin, 400 pmol of biotinylated peptides, up to 20 µg of biotinylated antibody, 20 µg of biotinylated double- stranded DNA, or 400 pmol of biotinylated single-stranded oligonucleotides.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.
For how long can the Streptavidin-coupled Dynabeads magnetic beads be stored?
Storage should be at 2 to 8 degrees C. Freezing or drying of the Dynabeads magnetic beads is not recommended. Provided the Dynabeads magnetic beads are stored correctly, quality is guaranteed until the expiry date stated on the label.
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How may I optimize binding capacity of streptavidin-coupled Dynabeads magnetic beads?
The binding capacity of streptavidin-coupled Dynabeads magnetic beads is fragment length-dependent. Reduced binding capacity for large DNA fragments may be due to steric hindrance. For large DNA fragments (greater than 2 kb in size), we recommend using Dynabeads kilobaseBINDER Kit.
-Salt concentration affects the binding efficiency of biotinylated nucleic acids to the Streptavidin-coupled Dynabeads magnetic beads. Optimal binding conditions for biotinylated DNA fragments (up to 1 kb) are achieved at 1 M NaCl (final concentration), 25 degrees C and 15 min incubation time. Longer DNA fragments should be immobilized overnight. Biotinylated antibodies should be immobilized in PBS buffer pH 7.4, supplemented with 0.1% BSA.
-Ensure that your sample does not contain an excess of free biotin, as the free biotin will bind Streptavidin-coupled Dynabeads magnetic beads much more rapidly than larger biotinylated molecules. Biotinylated oligonucleotides should be recovered by reverse phase HPLC or FPLC to remove free biotin from the sample.
-We also recommend a titration to optimize the quantity of beads used for each individual application, since both the size of the specific molecule to be immobilized and the biotinylation procedures affect the binding capacity of the beads.
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I have used biotinylated antibodies coupled to Dynabeads Streptavidin magnetic beads for immunoprecipitation (IP). How do I elute my target protein from the antibody without eluting the antibody off the beads?
Use mild elution conditions, e.g., a buffer with high salt or low pH. Heating the beads at 95 degrees C for 5 mins in SDS sample buffer will elute the antibody as well.
Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.