Product description
background | the rubisco quantitation kit is suitable for quantitation of Rubisco in plant and algal samples using quantitative immunoblotting. It contains anti-RbcL antibody, recombinant RbcL protein and the Agrisera PEB buffer for quantitative protein extraction at a price that is 20% lower as the individually purchased products. This kit is a good alternative to quantification of Rubisco by ELISA as it allows separation of Rubisco from a total complex cell protein mixture, which can be troublesome in case of ELISA. kit contains | ►rabbit polyclonal antibody to RbcL AS03 037, quantity | 100 µg amount enough for 100 Western blots ►Rubisco (RbcL) recombinant protein standard AS01 017S, quantity | 250 µl at 0.15 pmoles/µl, amount enough for generation of standard curve in 85 assays (standard curve: 0.0625 pmoles, 0.125 pmoles, 0.25 pmoles) ►Agrisera PEB extraction buffer AS08 300, quantity | 3 x 2 ml (4x stock), amount enough for 48 isolations of plant material (using 500 µl 1x PEB for 100 mg fresh weight) or 120 isolations of algal material (using 200 µl 1x PEB for cell amounts corresponding to 4-10 ug total chlorophyll) ►manual an article on quantitative immunoblotting can be found here. Kubien et al. (2011). Quantification of the amount and activity of Rubisco in leaves. Methods Mol Biol. 2011;684:349-62. application example
2 µg of total protein from various plant extracts (1-5) extracted with PEB (AS08 300) separated on 4-12% NuPage (Invitrogen) LDS-PAGE and blotted 1h to PVDF. Markers MagicMarks (Invitrogen) (M) and Rubisco protein standard (AS01 017S) at 0.0625 pmol, 0.125 pmol, 0.25 pmol. Following standard western blot procedure this image has been obtained using a CCD imager (FluorSMax, Bio-Rad) and Quantity One software (Bio-Rad). The contour tool of the software is used to select the area for quantitation and the values are background subtracted to give an adjusted volume in counts for each standard and sample.
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