Determination of amino acid compositions and NH2-terminal sequences of peptides electroblotted onto PVDF membranes from Tricine-Sodium dodecyl sulfate-polyacrylamide gel electrophoresis: Application to peptide mapping of human complement component C3

Decreased carbonic anhydrase III levels in the liver of the mouse mutant ‘toxic milk’ (tx) due to copper accumulation

“…Peptides were stacked at 30 V and electrophoresed at 130 V. After electrophoresis the gel was either stained with Coomassie Blue or equilibrated in 10 mM Caps, 20 % methanol, pH 11, for 10 min and transferred in the same buffer to an Immobilon membrane (Millipore) at 0.2 mA\cm for 2 h [19]. The membrane was stained with 0.1 % (w\v) Coomassie Blue and destained using 50 % aqueous methanol to identify the location of the peptides, and the excised bands were sequenced directly.…”

Purification and partial amino acid sequence of the glutamate 1-semialdehyde aminotransferase of barley and Synechococcus

“…Routinely the system described in (11) was used which contained 10% or 12.5% polyacrylamide. For the purpose of amino acid sequence determination proteins were separated in a Tricine-SDS-gel system (12,14). These gels were prepared with different polyacrylamide concentrations depending on the size of the peptides in the samples.…”

“…Gels were subjected to a pre-electrophoresis for 5 h at 30 mA with 20 mM thioglycolate in the upper reservoir buffer (12). Samples dissolved in 4% SDS, 12% glycerol, 50 mM Tris and 2% mercaptoethanol were heated for 30 min at 40 ~ and loaded on the gel.…”

Crystallisation and preliminary X‐ray analysis of the receptor‐binding domain of human and bovine α2‐macroglobulin

“…Amino acid analysis was performed as described [19], after hydrolysis with 6 M HCI, 0.1% phenol and 5% thioglycolic acid at 110 °C for 16 h. Excised bands from the ProBIott membrane was wetted with methanol prior to hydrolysis [20], The values of Ser and Thr were corrected for a loss of 10 and 5%, respectively. To determine the amount of aminosugar, hydrolysis was performed as described above, but only for 3 h. The values of GlcNH: was corrected for a loss of 5%.…”