C9891

Collagenase from Clostridium histolyticum

Type IA, 0.5-5.0 FALGPA units/mg solid, ≥125 CDU/mg solid, For general use

• 동의어(들): Clostridiopeptidase A
• CAS Number: 9001-12-1
• 효소 위원회 번호: 3.4.24.3 (BRENDA, IUBMB)
• EC Number: 232-582-9
• MDL number: MFCD00130830
• UNSPSC 코드: 12352204
• NACRES: NA.54

속성

• Quality Level: 200
• 양식: essentially salt-free, lyophilized powder
• 특이 활성도: ≥125 CDU/mg solid; 0.5-5.0 FALGPA units/mg solid
• 분자량: 68-130 kDa
• 기술: cell culture | mammalian: suitable; single cell analysis: suitable
• solubility: TESCA buffer (50 mM TES, 0.36 mM Calcium chloride, pH 7.4): soluble
• 응용 분야: diagnostic assay manufacturing
• 배송 상태: wet ice
• 저장 온도: −20°C

설명

애플리케이션

The enzyme has also been used along with other proteases for the disaggregation of human tumor, mouse kidney, human brain, and lung epithelial tissues. It is also useful in liver and kidney perfusion studies, digestion of pancreas, isolation of nonparenchymal rat liver cells and hepatocytes. The enzyme from Sigma has been used for the digestion of type I collagen from bovine trabecular and cortical bones. It has also been used as a positive control in FALGPA assay on Group B Streptococci cells.

Suitable for use in preparation of single cell suspension for sequencing.

생화학적/생리학적 작용

Collagenase is activated by four gram atom calcium per mole enzyme. It is inhibited by ethylene glycol-bis(beta-aminoethyl ether) - N, N, N′,N′-tetraacetic acid, beta-mercaptoethanol, glutathione, thioglycolic acid and 8-hydroxyquinoline.

Effective release of cells from tissue requires the action of both collagenase enzymes and the neutral protease. Collagenase is activated by four gram atom calcium (Ca²⁺) per mole enzyme. The culture filtrate is thought to contain at least 7 different proteases ranging in molecular weight from 68-130 kDa. The pH optimum ranges from 6.3-8.8. The enzyme is typically used to digest the connective components in tissue samples to liberate individual cells. Collagenase treatment can cause some cells to die. Typically, concentrations varying from 0.1 to 5 mg/mL are used for digestion. The duration of reaction varies from 15 minutes to several hours for a satisfactory efficiency of cell dissociation without causing too much cell death. Krebs Ringer buffer with calcium and BSA is preffered and Zn²⁺ is required for activity. The enzyme recognizes the sequence -R-Pro-8-X-Gly-Pro-R-, where X is most often a neutral amino acid. Ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA)4; β-mercaptoethanol; glutathione (reduced); thioglycolic acid, sodium; 2,2′-dipyridyl; 8-hydroxyquinoline, cysteine are known to inhibit the enzyme activity.

주의사항

As supplied, this product is stable for one year at -20°C. There is no loss in FALGPA or protease activity in 30 days at 37°C, 50°C and -20°C. Solutions of crude collagenase are stable if frozen quickly in aliquots (at 10 mg/mL) and kept frozen at -20°C. Further freeze-thaw cycles will damage the solution. The product retains 100% activity over 7 hours when held on ice.

단위 정의

One collagen digestion unit (CDU) liberates peptides from collagen from bovine achilles tendon equivalent in ninhydrin color to 1.0 μmole of leucine in 5 hours at pH 7.4 at 37 °C in the presence of calcium ions. One FALGPA hydrolysis unit hydrolyzes 1.0 μmole of furylacryloyl-Leu-Gly-Pro-Ala per min at 25°C. One Neutral Protease unit hydrolyzes casein to produce color equivalent to 1.0 μmole of tyrosine per 5 hr at pH 7.5 at 37°C. One Clostripain Unit hydrolyzes 1.0 μmole of BAEE per min at pH 7.6 at 25°C in the presence of DTT.

제조 메모

This product is equivalent to first 40% ammonium sulfate fraction of Mandl, I., et al., J. Clin. Invest., 32, 1323 (1953). Solutions are typically prepared at 1-2 mg/mL in TESCA buffer (containing 50 mM TES, 0.36 mM Calcium chloride, pH 7.4 at 37°C.

안전성 정보

• 픽토그램: GHS08,GHS07
• 신호어: Danger
• 유해 및 위험 성명서: H315,H319,H334,H335
• 예방조치 성명서: P261 - P264 - P271 - P280 - P302 + P352 - P305 + P351 + P338
• Hazard Classifications: Eye Irrit. 2 - Resp. Sens. 1 - Skin Irrit. 2 - STOT SE 3
• 표적 기관: Respiratory system
• Storage Class Code: 11 - Combustible Solids
• WGK: WGK 1
• Flash Point (°F): Not applicable
• Flash Point (°C): Not applicable
• 개인 보호 장비: dust mask type N95 (US), Eyeshields, Faceshields, Gloves