Alanine Transaminase Activity Assay Kit (Colorimetric/Fluorometric) (ab105134)

Overview

  • Product name
    Alanine Transaminase Activity Assay Kit (Colorimetric/Fluorometric)
  • Detection method
    Colorimetric/Fluorometric
  • Sample type
    Cell culture supernatant, Urine, Serum, Plasma, Other biological fluids, Tissue Extracts, Cell culture media
  • Assay type
    Enzyme activity (quantitative)
  • Sensitivity
    > 10 mU/well
  • Assay time
    1h 20m
  • Product overview

    Alanine Transaminase Activity Assay Kit (Colorimetric/Fluorometric) ab105134 is a rapid and simple assay used to quantify alanine transaminase (ALT) activity in mammalian samples.


    In the ALT assay protocol, ALT transfers an amino group from alanine to α-ketoglutarate; producing pyruvate and glutamate. The pyruvate is detected in a reaction that converts a nearly colorless probe to a form that is colored (ODmax = 570 nm) and fluorescent (Ex/Em = 535/587 nm).


    The kit has a detection limit of 10 mU per well.


    ALT assay protocol summary:
    - add samples and standards to wells
    - add reaction mix and incubate for 10 min at 37ºC
    - analyze every 2-3 min for 60 min with microplate reader in kinetic mode at 37ºC

  • Notes

    Alanine transaminase is also called alanine aminotransferase or serum glutamic pyruvic transaminase (ALT, ALAT, SGPT).

  • Platform
    Microplate reader

Properties

Images

  • Serum aspartate transaminase (AST) and alanine transaminase (ALT) levels were measured at various time periods post Con A injection. Mean values ± SD are shown (n = 4). ∗P < 0.05 and ∗∗P < 0.01.
  • Liver samples from high fat diet (HFD) and standard carbohydrate diet (CHD) BALB/c and C57BL6/J mice were homogenised in an ALT assay buffer for the determination of ALT activity using ab105134. A separate batch of liver extracts was prepared and incubated in a buffer containing NP40 (5%) and supernatants containing the triglycerides were separated. Triglycerides concentration was determined on the supernatant fraction using ab65336. ALT activity and triglycerides concentration were determined by measuring OD at 570nm.

  • Colorimetric standard curve: mean of duplicates (+/- SD) with background reads subtracted.
  • Fluorometric standard curve: mean of duplicates (+/- SD) with background reads subtracted.

  • Alanine transaminase measured in mouse tissue lysates showing quantity (mU) per mg of tested sample.

    Protein concentration for samples varied from 4 mg/mL to 13 mg/mL. Samples were diluted 9-27 fold and measured colorimetrically.

  • Pyruvate measured colorimetrically in cell lysate after 20 min and 40 min incubation time showing quantity (nmol) per 1 mln of tested cells.

  • Measurement of alanine transaminase in HepG2 cells (10 μg) and liver lysate (15 μg).

  • Pyruvate measured fluorometrically in cell lysate after 20 min and 40 min incubation time showing quantity (nmol) per 1 mln of tested cells.

  • Pyruvate measured in biological fluids after 20 min and 40 min incubation time showing quantity (nmol) per ml of tested sample.

  • Pyruvate measured in mouse tissue lysates after 20 min and 40 min incubation time showing quantity (nmol) per mg of tested sample.

Protocols

References

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Submitted Jan 24 2018

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