Wako Product Name

HR Series NEFA-HR(2)

For In Vitro Diagnostic Use

Special Characteristics

  • Detects a variety of Free Fatty Acids
    Serum NEFA Recovery Study
    Free Fatty Acid (NEFA) Activity % Recovery %
    Acetic acid00
    Proprionic acid00
    Butyric acid00
    Valeric acid149
    Caproic acid9899
    Caprylic acid9794
    Capric acid8986
    Lauric acid9897
    Myristic acid10393
    Palmitic acid9794
    Stearic acid90-
    Arachidic acid94-
    Palmitoleic acid100103
    Oleic acid100102
    Linoleic acid99102
    Linolenic acid102100
    Arachidonic acid8985
    Citric acid00
    Oxalic acid00
  • Minimal interferences from other serum components such as uric acid, glucose, hemoglobin, and Bilirubin.
  • Specific to Free Fatty Acids with no interferences from other related compounds.

Extraction methods are widely used for the colorimetric determination of non-esterified fatty acids (NEFA) in serum. NEFA are converted to their copper salts that are extracted into an organic solvent. The salts are then complexed with a dye for purposed of colorimetric measurement. Alternatively, extracted NEFA are titrated with standard alkali to an acid base indicator endpoint. These approaches are time consuming, hazardous and not easily automated. Wako has made extensive studies of NEFA quantitation and has succeeded in developing an original enzymatic method which is available in a series of individual reagents. This enzymatic method is accurate, precise, simple and fast. The need for an extraction step has been eliminated and the method can be automated.

The Wako enzymatic method relies upon the acylation of coenzyme A (CoA) by the fatty acids in the presence of added acyl-CoA synthetase (ACS). The acyl-CoA thus produced is oxidized by added acyl-CoA oxidase (ACOD) with generation of hydrogen peroxide, in the presence of peroxidase (POD) permits the oxidative condensation of 3-methy-N-ethyl-N(β-hydroxyethyl)-aniline (MEFA) with 4-aminoantipyrine to form a purple colored adduct which can be measured colorimetrically at 550 nm.

Intended Use The Wako HR series NEFA-HR(2) is an in vitro enzymatic colorimetric method assay for the quantitative determination of non-esterified fatty acids (NEFA) in serum.
Medical Conditions
Method Enzymatic
Product Brochure Wako NEFA Brochure
Assay Linearity 0.01-4.00 mEq/L
Basic Procedure Sample: 4 µL
R1: 225 µL
R2: 75 µL
Wavelength: primary: 560 nm, secondary: 670 nm
Package Insert HR Series NEFA-HR(2)
  • To request an automated analyzer application, please email Please include the names of the product and the analyzer/instrument.

Ordering Information

Catalog No. Product Name Pkg Size Storage
999-34691 HR Series NEFA-HR(2) Color Reagent A 4 x for 50 mL 2-10°C
995-34791 HR Series NEFA-HR(2) Solvent A 4 x 50 mL 2-10°C
991-34891 HR Series NEFA-HR(2) Color Reagent B 4 x for 25 mL 2-10°C
993-35191 HR Series NEFA-HR(2) Solvent B 4 x 25 mL 2-10°C


276-76491 NEFA Standard Solution    

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997-76491 Wako NEFA Linearity Set