Alanine Transaminase Assay Kit

Produktnummer ABIN1000289

Produktdetails

Target: ALT (Alanine Aminotransferase (ALT))

Applikation: Activity Assay (AcA)

Proben: Plasma, Serum

Spezifität: 2 U/L

Produktmerkmale: Sensitive. Linear detection range: 2-100 U/L.
Simple and convenient. This simple, convenient assay can be carried out in a microplate or a cuvette and takes only 10 min.

Bestandteile: Assay Buffer: 24 mL. LDH: 120 µL. Cosubstrate: 600 µL. NADH: 500 µL.

Benötigtes Material: Pipeting devices and accessories. Clear bottom 96-well plates (e.g. Corning Costar) and plate reader or spectrophotometer and cuvettes for measuring OD340nm.

Anwendungsinformationen

Applikationshinweise: Direct Assays: ALT activity in serum, plasma and other biological samples.
Drug Discovery/Pharmacology: effects of drugs on ALT activity.

Protokoll: Procedure using 96-well plate
1.Samples and controls. Transfer 20 µL of each sample to separate wells. Also, for each assay plate, include two wells with 20 µL distilled water to be used for the NADH Standard and Blank. Keep plate at the desired temperature (e.g. 37°C).
2.Prepare Working Reagent for Sample and Standard wells by mixing for each well: 200 µL Assay Buffer, 5 µL Cosubstrate, 1 µL LDH and 4 µL NADH. Warm to desired temperature (e.g. 37°C). Prepare Blank Reagent for the Blank well by mixing: 200 µL Assay Buffer, 5 µL Cosubstrate, 1 µL LDH and 4 µL H2O. Warm to desired temperature (e.g. 37°C).
3. Add 200 µL Working Reagent to each Sample and Standard well, and 200 µL Blank Reagent to the Blank well. Immediately tap plate to mix, incubate at the desired temperature and read OD340nm at 5 min and at 10 min. Alternatively, record kinetics at 340 nm.

Procedure using cuvettes
1. For each assay, include one Standard and one Blank control. For each Sample and Standard, prepare Working Reagent by mixing 1000 µL Assay Buffer, 25 µL Cosubstrate, 5 µL Enzyme Mix and 20 µL NADH. Transfer 1000 µL Working Reagent to each sample cuvette and standard cuvette. Warm to desired temperature (e.g. 37°C). For Blank control, mix 1000 µL Assay Buffer, 25 µL Cosubstrate, 5 µL Enzyme Mix and 20 µL H2O. Transfer 1000 µL Reagent to the Blank control cuvette. Warm to desired temperature (e.g. 37°C).
2. Prewarm sample to the desired temperature. Add 100 µL Sample to the Sample Cuvette. Transfer 100 µL H2O to the Standard cuvette and to Blank Control cuvette, respectively. Mix immediately.
3. Read OD340nm at 5 min and 10 min. Alternatively, record kinetics at 340 nm.

Aufbereitung der Reagenzien:

Equilibrate all components to room temperature. Mix assay buffer well by vigorous shaking. Keep thawed enzyme on ice. Assays can be performed at 37°C or at room temperature. Prior to assay, bring the working reagents, microplate and spectrophotometer to the desired temperature. Assay is compatible with serum or plasma (heparin, EDTA). Samples should be clear and free of particles or precipitates. Hemolyzed samples should not be used.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Lagerung: -20 °C

Referenzen

Dankel, Degerud, Borkowski, Fjære, Midtbø, Haugen, Solsvik, Lavigne, Liaset, Sagen, Kristiansen, Mellgren, Madsen: "Weight cycling promotes fat gain and altered clock gene expression in adipose tissue in C57BL/6J mice." in: American journal of physiology. Endocrinology and metabolism, Vol. 306, Issue 2, pp. E210-24, (2014) (PubMed).

Kim, Joe, Kong, Jeong, Cho, Ryter, Chung: "Carbon monoxide protects against hepatic ischemia/reperfusion injury via ROS-dependent Akt signaling and inhibition of glycogen synthase kinase 3β." in: Oxidative medicine and cellular longevity, Vol. 2013, pp. 306421, (2014) (PubMed).

Peng, Xu, Zhang, Zhang, Huang, Fu, Xue, Da, Dai, Qiao, Dong, Zhang, Meng: "Thermosensitive injectable hydrogel enhances the antitumor effect of embelin in mouse hepatocellular carcinoma." in: Journal of pharmaceutical sciences, Vol. 103, Issue 3, pp. 965-73, (2014) (PubMed).

Mani, Ramasamy, Ahmad, Wahab, Jaafar, Kek, Salleh, Majeed: "Effects of the total alkaloidal extract of Murraya koenigii leaf on oxidative stress and cholinergic transmission in aged mice." in: Phytotherapy research : PTR, Vol. 27, Issue 1, pp. 46-53, (2013) (PubMed).

Zhang, Xu, Weihrauch, Jones, Jing, Shi, Gourlay, Oldham, Hillery, Pritchard: "Inhibition of myeloperoxidase decreases vascular oxidative stress and increases vasodilatation in sickle cell disease mice." in: Journal of lipid research, Vol. 54, Issue 11, pp. 3009-15, (2013) (PubMed).

Rattan, Graham, Maguire, Giri, Shridhar: "Metformin suppresses ovarian cancer growth and metastasis with enhancement of cisplatin cytotoxicity in vivo." in: Neoplasia (New York, N.Y.), Vol. 13, Issue 5, pp. 483-91, (2011) (PubMed).

Antigendetails

Target: ALT (Alanine Aminotransferase (ALT))

Andere Bezeichnung: Alanine Transaminase (ALT Produkte)

Synonyme: AAT1 Kit, ALT1 Kit, GPT1 Kit, GPT Kit, Gpt1 Kit, An11g02620 Kit, AO090003000164 Kit, 233.t00009 Kit, 24.t00016 Kit, 1300007J06Rik Kit, 2310022B03Rik Kit, ALT Kit, Gpt-1 Kit, AU021132 Kit, Dpagt2 Kit, Gnpta Kit, gpt Kit, ALANINE AMINOTRANSFERAS Kit, AtAlaAT1 Kit, AtAlaATc Kit, T13M22.3 Kit, T13M22_3 Kit, alanine aminotransferas Kit, glutamic--pyruvic transaminase Kit, glutamic-pyruvate transaminase (alanine aminotransferase) Kit, alanine aminotransferase Kit, alanine aminotransferase 1 Kit, glutamic pyruvic transaminase, soluble Kit, dolichyl-phosphate (UDP-N-acetylglucosamine) acetylglucosaminephosphotransferase 1 (GlcNAc-1-P transferase) Kit, alanine aminotransferase 2-like Kit, Alanine aminotransferase 1, mitochondrial Kit, GPT Kit, Gpt Kit, Tb927.1.3950 Kit, ANI_1_368094 Kit, AOR_1_284154 Kit, ALAT_1 Kit, ALAAT1 Kit, EHI_159710 Kit, EHI_096750 Kit, LOC5572541 Kit, ALAT Kit, AAT2 Kit, Dpagt1 Kit, LOC100537633 Kit, AlaAT1 Kit

Hintergrund: Quantitative determination of Alanine transaminase activity at 340nm.
Procedure: 10 min.

Alanine Transaminase (ALT), also known as serum alanine aminotransferase (ALAT) or pyruvic transaminase (SGPT), facilitates the conversion of alanine and -ketoglutarate to pyruvate and glutamate. ALT plays an important role in gluconeogenesis and amino acid metabolism. ALT is found mainly in the liver, and, to a lesser extent, in kidney, heart, muscle, and pancreas tissues. Normal serum levels of ALT are low, and increased serum ALT activity is widely used as a marker for liver damage. Simple, direct and automation-ready procedures for measuring ALT activity find wide applications in research and drug discovery. This ALT activity assay is based on the quantification of pyruvate produced by ALT. In this assay, pyruvate and NADH are converted to lactate and NAD by the enzyme lactate dehydrogenase (LDH). The decrease in NADH absorbance at 340 nm is proportional to ALT activity.