KEY FEATURES

Sensitive and accurate. Use as little as 10 μL samples. Linear detection range 0.01 mmol/L to 1.0 mmol/L (0.88 mg/dL to 88.5 mg/dL) triglyceride.

APPLICATIONS:

Direct Assays: triglyceride in biological samples (e.g. serum and plasma).

KIT CONTENTS

Assay Buffer: 24 mL ATP: 250 μL Dye Reagent: 220 μL

Enzyme Mix: 500 μL Lipase: 1000 μL

Standard: 100 μL (equivalent to 100 mmol/L Triglyceride)

Storage conditions. The kit is shipped on dry ice. Store Assay Buffer at 4°C and other reagents at -20°C. Shelf life of three months after receipt.

Precautions: reagents are for research use only. Normal precautions for laboratory reagents should be exercised while using the reagents. Please refer to Material Safety Data Sheet for detailed information.

Note: (1) SH-group containing reagents (e.g. mercaptoethanol, DTT) may interfere with this assay and should be avoided in sample preparation; (2) if sample contains glycerol, use BioAssay Systems' EnzyChromTM Glycerol Assay Kit (Cat# EGLY-200) to determine glycerol concentration and subtract the glycerol value to yield triglyceride concentration.

1. Equilibrate all components to room temperature. Keep thawed Lipase and Enzyme Mix in a refrigerator or on ice. Dilute Standard in distilled water as follows. Transfer 10 μL diluted standards into wells of a clear 96-well plate. Diluted standards can be used for future assays when stored refrigerated.

Serum and plasma samples should be diluted 5-fold in dH2O and are assayed directly. Cells and other solid samples can be solubilized in 5% Triton X-100 (see Ref. 3). Transfer 10 μL samples into separate wells of the 96-well plate.

2. Prepare Working Reagent for each well, by mixing 100 μL Assay Buffer, 2 μL Enzyme Mix, 5 μL Lipase, 1 μL ATP and 1 μL Dye Reagent in a clean tube. Transfer 100 μL Working Reagent into standards and sample wells. Tap plate to mix.

3. Incubate 30 min at room temperature. Read optical density at 570nm (550-585nm).

Note: 1. if the Sample OD is higher than the Standard OD at 1.0 mmole/L triglyceride, dilute sample in water and repeat the assay. Multiply by the dilution factor n.

Subtract ODH2O (water, #4) from the standard OD values and plot the OD against standard concentrations. Determine the slope using linear regression fitting. The triglyceride concentration of Sample is calculated as

ODSAMPLE and ODH2O are optical density values of the sample and the water blank (# 4). n is the dilution factor. For example serum or plasma samples are diluted 5-fold prior to assay, n = 5.

Conversions: 1 mmol/L triglyceride equals 88.5 mg/dL or 10 ppm.