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The data we get from reading abcam dipsticks used on lysed whole blood has 5-6 peaks. We would like to know what each peak refers to and which to consider when analyzing data. Is it necessary to change any settings on the reader when reading dipsticks from different assay kits? Can you provide us with an example of analyzed data obtained from the dipstick reader? The dipstick kits we are having trouble with are… ab109720 – Complex I Enzyme Activity Dipstick Assay Kit ab109876 – Complex IV Enzyme Activity Dipstick Assay Kit Thank you,
Asked on Feb 17 2012
Thank you for contacting Abcam regarding this issue.
On occasion the reader will see more profiles which cannot be seeing with the naked eye on the dipstick. This is due sometimes to imperfections of the nitrocellulose or to uneven wicking. For activity assays the antibody line is stripped at about 7mm from the nitrocellulose edge of the dipstick (or lower end of the dipstick). In new readers, the dipstick is inserted with the wicking pad towards the door of the instrument and the nitrocellulose towards the inside of the instrument. The reader will read from the inside of the instrument (lower section of the dipstick) to the outside of the instrument (upper section of the dipstick). Therefore the lower section of the dipstick will appear towards the left of the absorption profile. The reader will pick up all peaks in a sample .
We strongly recommend creating a standard curve using 7 to 8 dipsticks for covering the working range. In this way you will be able to know which peaks are background and which are signal. Following the protein concentration ranges as defined in Table 1 of the protocol, generate a standard curve using a positive control sample. We recommend performing a 1:2 serial dilution with 1 part Buffer A to 1 part Buffer B in a total volume of 50 µl.
Complex IV dipsticks may sometimes develop a percipitate about 2mm from the edge, this should be ignored.
I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.
Answered on Feb 17 2012