Peptides from a sample that are not captured during the loading step
The stationary phase used in our columns has a lower retentivity of peptides than other stationary phases (peptides are eluted at a lower %ACN than other stationary phases).
Therefore, there is a small amount of peptides that will not bind to the stationary phase, even under no or low %ACN conditions. Generally, the percentage of unbound peptides represents a small percentage of the peptides from the sample.
The amount of peptides that escape capture during loading can be minimised by loading at 0% or 1% ACN.
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Sample loading
We recommend the following buffer compositions: Buffer A: 99.9% MilliQ Water, 0.1% formic acid Buffer B: 99.9% Acetonitrile, 0.1% formic acid Please consult with your UHPLC manufacturer to confirm that your UHPLC is compatible with these compositions ...
Sample preparation
Prior to loading samples onto your Aurora Series column, the sample should be de-salted and should not contain any contaminants (salts, detergents, solid particles, etc). Important Loading contaminated samples onto the column may disrupt solvent flow ...
Expected phosphopeptide identifications using Aurora Series columns
Counts shown on the Y-axis below represent the number of unique phosphorylated peptides identified across 4 replicate runs. These results were achieved under the following conditions: Column: 25cm Aurora Ultimate column Sample: 500ng ...
How to revive a column that has been blocked by a 'thick' sample
A thick sample is often the result of precipitate forming after the addition of loading buffer to a sample vial. There are two things you can try to revive the column: Run 5 - 10+ blank gradients Run 85% acetonitrile at 400nl/min for an extended ...
Initial operation and buffer compositions
Once the column is connected to your UHPLC system and is placed inside the source heater or housing, begin operation using 70% buffer B at a flow rate equivalent to the desired gradient flow rate for around 10 minutes or until the pressure is stable ...