title: Amino acid analysis.
abstract: Amino acid analysis (AAA) is one of the best methods to quantify peptides and proteins. Two general approaches to quantitative AAA exist, namely, classical postcolumn derivatization following ion-exchange chromatography and precolumn derivatization followed by reversed-phase HPLC (RP-HPLC). Excellent instrumentation and several specific methodologies are available for both approaches, and both have advantages and disadvantages. This unit focuses on picomole-level AAA of peptides and proteins using the most popular precolumn-derivatization method, namely, phenylthiocarbamyl amino acid analysis (PTC-AAA). It is directed primarily toward those interested in establishing the technology with a modest budget. PTC derivatization and analysis conditions are described, and support and alternate protocols describe additional techniques necessary or useful for most any AAA method--e.g., sample preparation, hydrolysis, instrument calibration, data interpretation, and analysis of difficult or unusual residues such as cysteine, tryptophan, phosphoamino acids, and hydroxyproline.
authors.affiliation: W. Alton Jones Cell Science Center, Lake Placid, New York, USA.
authors.forename: J W
authors.initials: JW
authors.lastname: Crabb
authors.forename: K A
authors.initials: KA
authors.lastname: West
authors.forename: W S
authors.initials: WS
authors.lastname: Dodson
authors.forename: J D
authors.initials: JD
authors.lastname: Hulmes
mesh_terms: D000096:Acetone; D000596:Amino Acids / Q000032:analysis*; D011232:Chemical Precipitation; D002851:Chromatography, High Pressure Liquid; D003956:Dialysis; D004986:Ether; D006868:Hydrolysis; D006909:Hydroxyproline / Q000378:metabolism; D008567:Membranes, Artificial; D008698:Mesylates / Q000378:metabolism; D008872:Microwaves; D010670:Phenylthiourea / Q000378:metabolism; D010750:Phosphoproteins / Q000032:analysis; D011145:Polyvinyls; D011506:Proteins / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism; D005465:Fluorocarbon Polymers
chemical_list: D000096:Acetone; D000596:Amino Acids; D004986:Ether; D006909:Hydroxyproline; D008567:Membranes, Artificial; D008698:Mesylates; D010670:Phenylthiourea; D010750:Phosphoproteins; D011145:Polyvinyls; D011506:Proteins; D013438:Sulfhydryl Compounds; C013976:dodecylmercaptan; C045880:methanesulfonic acid; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers