This document introduces the N-terminal 4.0 biphasic column protein sequencer methods of HP 241 protein sequencer, including N-terminal protein column 4.0, N-terminal protein column 4.0 (cycle 1), N-terminal peptide column 4.0, N-terminal peptide column 4.0 (cycle 1), N-terminal flask 4.0, PTH-Std 4.0, column prep 4.0 methods
This document describes the N-terminal 4.0 biphasic column sequencing methods implemented on the Hewlett-Packard HP 241 Protein Sequencer (N+C). These methods utilize Edman degradation chemistry to analyze protein and peptide samples in the low nanomole to low picomole range loaded onto a specific Hewlett-Packard biphasic RP/SAX column. The document outlines that reagents 1R, 2R, 3R, and TH-Std are specific for C-terminal sequencing and are not utilized here.
The text provides detailed step-by-step instructions for several methods, including N-Terminal Protein Column 4.0, N-Terminal Peptide Column 4.0, N-Terminal Flask 4.0, PTH-Std 4.0, and Column Prep 4.0. It specifies the required bottle configuration of reagents and solvents purchased from Hewlett-Packard, such as Diphenylphosphorylisothiocyanate (DPP-ITC) and various buffers. The document also details the specific steps for coupling, cleavage, conversion, washing, and extraction cycles with associated times and temperatures.
| Sample Range | low nanomole to low picomole range |
|---|---|
| Waste Volume | less than 1 mL/cycle |
| Reagent Source | Hewlett-Packard |
It provides a longer initial drying time and a pause for proper HPLC column equilibration after switching from C-terminal sequencing.
Reagents 1R, 2R, 3R, and TH-Std are not utilized; the document focuses on methods using specific coupling agents like DPP-ITC.