Hi, We have 10ul of a low quantity library (700pM) that we are trying to denature and load onto the flow cell at 10pM.
So far, the best that I can come up with while still keeping the NaOH concentration low enough is 9.5 ul of the sample + 0.5 ul of 2N NaOH and load all 10ul in 990ul of HT1, giving a final pM concentration of 6.65. Unfortunately this won't give us the cluster density that we would like. I could adjust the HT1 volume but that would increase the NaOH and probably inhibit clustering somewhat, if not all together. Could I double the incubation time and decrease the NaOH concentration by half? I'm not sure if the denaturation would work at half the normal concentration. Does anyone have another solution? Any suggestions would be really appreciated.
So far, the best that I can come up with while still keeping the NaOH concentration low enough is 9.5 ul of the sample + 0.5 ul of 2N NaOH and load all 10ul in 990ul of HT1, giving a final pM concentration of 6.65. Unfortunately this won't give us the cluster density that we would like. I could adjust the HT1 volume but that would increase the NaOH and probably inhibit clustering somewhat, if not all together. Could I double the incubation time and decrease the NaOH concentration by half? I'm not sure if the denaturation would work at half the normal concentration. Does anyone have another solution? Any suggestions would be really appreciated.
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