![]() ![]() ![]() Following the incubation, remove the antibody solution and wash three more times with 1 x TBST IX) After washing, incubate the membrane with secondary antibody which is conjugated with HRP (Horse Radish Peroxidase) and diluted in 01xTBS with 0.5% BSA for 1 hr at room temperature. As with a western blot, samples are separated by electrophoresis and transferred onto a membrane. It has been applied to detect Restriction Fragment Length Polymorphism (RFLP) and Variable Number of Tandem Repeat (VNTR) Polymorphism. The method is named after its inventor, the British biologist Edwin Southern. Once the incubation time is over, remove the primary antibody and wash the membrane three times with 1 x TBST Whereas a western blot is designed to detect specific proteins, a Southern blot, 2 developed in 1975 and named after its inventor Edwin Southern, 20 is used to detect specific DNA sequences. Southern blot is a method used to check for the presence of a DNA sequence in a DNA sample. The antibody should be diluted with 1xTBS with 0.5% BSA (w/v) according to the supplier’s instructions. The technique was named after its inventor, Edward Southern. VIII) After blocking, wash once with 1xTBS containing Tween 20 and incubate with primary antibody overnight at 4ᵒC. A Southern blot is a laboratory method used to detect specific DNA molecules from among a many other DNA molecules. Block the non-specific binding by incubating the membrane with 1xTBS containing 2% BSA w/v for 1 h at room temperature. VII) Wash the membrane with distilled water. WB: Western Blot SB: Southern Blot NB: Northern Blot PS: Protein Sequencing AA: Amino Acid Analysis. Place the memberane in a tray containing Ponceau S for staining. The sensitivity of the system is irrelevant if the signal cannot be distinguished from the noise. Optimizing the S/N ratio is often more important than increasing the sensitivity of the system. Disassemble the transfer apparatus from the top to downward, peeling off each layer in turn. In western blotting, the signal is the density of the specific protein band being probed for the noise is the density of the background. VI) After 7 min turn off the power supply and disconnect the leads. ![]()
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