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I used your urinary prostacyclin ELISA kit(ab133032) for measurement of urinary prostacyclin, in sample handling I have some problem, looking forward to your kind help.
1. How long time it will take for hexane pass through from the C18 reverse column ? (in my procedure hexane couldn’t pass from the column, is there something can effect column working?)
2. In protocol mention that when apply sample to the column the flow rate will be 0.5ml/min, what about another reagent like ethanol, deionized water and hexane?
3. For 1ml urine sample what volume coulmn we should use?
Asked on Feb 26 2013
The length of time could potentially vary depending on your lab set up. The person who optimized this extraction did not note the exact flow through time so unfortunately I cannot give you an exact recount of how long it took hexane to flow through for us. The flow through rate could depend again on lab set up for the extraction but also the size of column used. The size of column used could also vary depending on the amount of sample you have to run. For a more accurate indication of flow through rates and column size options (most columns have a max volume/ug of protein load) for your sample you will be best served to contact the column manufacturer directly. Below is the link for the column manufacturer we use for C18 columns and we often use cat#9002. They should be able to give you more specific information as well.
Answered on Feb 26 2013