Technical replicates (n=4) were averaged and plotted to determine the linear regression for the entire concentration range (R2=0.9994). The densitometric signal intensity was determined for each lane. Densitometric Signal Linearity versus Protein Load. As the SeeBlue Plus2 Pre-stained Protein Standard is already labeled, its protein bands are not all susceptible to labeling with No-Stain Reagent, and therefore are not all visible in this image.ī. c) ab116029 for smaller proteins down to 3.5 kDa. Two orange reference bands at 70 kDa and 25 kDa and one green reference band at 10 kDa highlight the blue-stained protein ladder. b) ab116028 for larger proteins up to 245 kDa. The PageRuler Plus Prestained Protein Ladder is a mixture of nine (9) blue-, orange- and green-stained proteins (10 to 250 kDa) for use in protein electrophoresis (SDS-PAGE) and western blotting. For most researchers, we generally recommend: a) ab116027 for proteins between 10 and 180 kDa. plus Western blotting detection system (Amersham, GE Healthcare) according to. When choosing the right pre-stained protein ladder, the most important choice is based on the size of your protein of interest. The image was captured using an iBright Imager with the No-Stain Membrane epi setting (455-485 nm excitation and 565-615 nm emission). SeeBlue (Invitrogen) protein ladder is included to allow molecular weights. The protein standard is supplied in a ready-to-use format for direct loading. The standards consist of three blue-dyed and two colored bands ranging in size from 21261 kDa for monitoring gel and transfer runs. After transfer, the proteins on the membrane were labeled following the No-Stain Protein Labeling Reagent protocol for midi-sized membranes. E-PAGE SeeBlue Pre-stained Standards are designed for use with the pre-cast E-PAGE Electrophoresis System. Proteins from the gel were transferred onto a 0.45 µm PVDF membranes at 25V constant for 30 minutes using the SureLock Tandem Midi Blot Module. The gel was electrophoresed using MES running buffer in the SureLock Tandem Midi Gel Tank. coli Lysate ranging from 2 to 0.25 µg and SeeBlue Plus2 Pre-stained Protein Standard in lanes 1-2 and 19-20. A NuPAGE 4-12% Bis-Tris Midi gel was loaded with recurring dilutions of E. Prestained protein ladders are recommended for. Uniform transfer using SureLock Tandem Midi Gel Tank. Total protein normalization of transferred proteins shows uniform transfer across the blot.Ī.
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