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Titolo:
NOVEL PROCEDURES FOR ISOLATING INTACT RETINAL VASCULAR BEDS FROM DIABETIC HUMANS AND ANIMAL-MODELS
Autore:
LAVER NM; ROBISON WG; PFEFFER BA;
Indirizzi:
NEI,BLDG 6,RM 316,9000 ROCKVILLE PIKE BETHESDA MD 20892 NEI,BLDG 6,RM 316,9000 ROCKVILLE PIKE BETHESDA MD 20892
Titolo Testata:
Investigative ophthalmology & visual science
fascicolo: 6, volume: 34, anno: 1993,
pagine: 2097 - 2104
SICI:
0146-0404(1993)34:6<2097:NPFIIR>2.0.ZU;2-9
Fonte:
ISI
Lingua:
ENG
Soggetto:
CELLS; RAT;
Keywords:
RETINA; CAPILLARIES; DIABETES; ELASTASE; TRYPSIN DIGESTION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
30
Recensione:
Indirizzi per estratti:
Citazione:
N.M. Laver et al., "NOVEL PROCEDURES FOR ISOLATING INTACT RETINAL VASCULAR BEDS FROM DIABETIC HUMANS AND ANIMAL-MODELS", Investigative ophthalmology & visual science, 34(6), 1993, pp. 2097-2104

Abstract

Purpose. To improve the 30-year-old ''trypsin digestion'' procedure for isolation of the complete retinal vasculature, which, in its time, was a revolutionary advance that allowed important discoveries about diabetic retinopathy; to provide a method that will yield more consistent results when applied to retinas representing a wide range of ages, species, and severity of vascular disease, such as that occurring in diabetes. Methods. Because the Difco trypsin preparation (Difco Laboratories, Detroit, MI) is a crude pancreatic extract, containing variableamounts of chymotrypsin, elastase, amylase, lipase, ribonuclease, collagenase, and other contaminants, an attempt was made to determine which of the major enzymes alone (using purified preparations), or what combination of enzymes, might be most effective in providing consistently clean yet intact retinal vasculatures from eyes of different origins. Results. Purified elastase alone (40 U/ml) in 100 mmol/l sodium phosphate buffer with 150 mmol/l sodium chloride and 5 mmol/l EDTA at pH 6.5 and 37-degrees-C gave better results than various concentrations of purified trypsin or chymotrypsin alone, or mixtures of trypsin/chymotrypsin, trypsin/elastase, chymotrypsin/elastase, or the crude trypsinpreparation. Conclusions. Elastase, which exhibits broad protease activity, and not trypsin, is the most important enzyme of the standard, crude trypsin digestion procedure for removal of the nonvascular tissues of the retina.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 06/12/20 alle ore 00:00:57