Lactate assay kit

Glucose Uptake Assay Kit (KA4086) and Lactate Assay Kit (KA0833) were obtained from Abnova (Taiwan, China). Basing on the kit instructions, the glucose uptake and lactate production of HSC6 and OECM1 cells were determined.

MCDB-131 cell culture medium was obtained from Gibco (Paisley, Scotland, UK). Other cell media, penicillin, streptomycin and amphotericin B, and trypsin were purchased from BioWhittaker (Verviers, Belgium). Fetal bovine serum (FBS) was purchased from Biowest (Kansas, USA). 2-NBDG (2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose) and BODIPY FL C₁₆ (4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-hexadecanoic acid) were supplied by Molecular Probes (Eugene, OR, USA). L-[¹⁴C(U)]-Glutamine was acquired from Perkin Elmer (Waltham, Massachusetts, USA). Lactate assay kit was from Abnova (Taoyuan City, Taiwan). Anti-MCT1 antibody was from Santa Cruz Biotechnology (Heidelberg, Germany), anti-HIF-1α antibody was from BD Biosciences (Franklin Lakes, NJ, USA) and anti-LC3B and anti-α-tubulin antibodies were from Cell Signaling Technology (Danvers, MA, USA). Plastic material for cell culture was from Nunc (Roskilde, Denmark). All other reagents not listed on this section, including sodium palmitate, were from Sigma-Aldrich (St. Louis, MO, USA).

Glucose and lactate concentrations were measured from supernatants using a colorimetric assay. The Glucose Assay Kit and the Lactate Assay Kit were purchased from Abnova (KA0831 and KA0833) and used according to the manufacturer’s recommendations. To fit into the linear concentration range of the assay, samples were diluted at 1:200 for glucose assay and 1:50 for lactate assay. Data were acquired using a Fluostar OPTIMA BMG Labtech plate reader and analyzed with the optima data analysis software (version 3.32).