Protein Reagents – Total Protein (Biuret Method)

Intended Use: This reagent kit is used for in-vitro quantitative determination of Total Protein in serum and plasma. 

Pack size: 2 × 50 ml 

Shelf life: 18 months 

Storage: 2° to 8° C 

Product overview

VDx Total Protein (Biuret Method) is a liquid reagent for the quantitative determination of total protein in serum or plasma. In an alkaline medium, peptide bonds react with cupric ions in Biuret reagent to form a violet-coloured complex with an absorbance maximum at 546 nm (530–570 nm). Colour intensity is directly proportional to total protein concentration in the sample. 

For general background on what a total protein test measures (albumin + globulin) and how it’s used clinically, see:

MedlinePlus: Total protein test

MedlinePlus: Total protein and A/G ratio

Key features 

• Method: Biuret, end-point 
• Wavelength: 546 nm (530–570 nm) 
• Linearity: 10 gm/dl 
• Reaction time (manual): 20 minutes at 21–25°C or 5 minutes at 37°C 
• Interference (IFU note): reagent is free of interference from moderate levels of haemoglobin, bilirubin, lipids, and salicylates; avoid grossly haemolysed samples or lipaemic sera 

Intended use

This reagent kit is used for in-vitro quantitative determination of Total Protein in serum and plasma. 

Test principle

In alkaline medium, peptide bonds of proteins react with cupric ions in Biuret reagent to form a violet-coloured complex with absorbance maximum at 546 nm (530–570 nm). Intensity of colour formed is directly proportional to the concentration of total proteins in the sample. 

Kit contents

• Reagent 1: Biuret Reagent
• Reagent 2: Calibrator (concentration as indicated on the label)
• Insert: 01 No. 

Specimen collection and storage

• Specimen: serum or heparinised plasma 
• Samples should preferably be used on the same day. If necessary, samples may be preserved up to one week if stored at 2–8°C.
• Bring sample to room temperature prior to use. 

Storage and stability

• VDx Total Protein liquid reagents are stable until the expiry date indicated on the labels when stored at 2–8°C.
• Working reagent stability: six months at room temperature. 

Manual procedure 

Pipette into test tube: 

• Biuret reagent: 1 ml (Blank / Standard / Test)
• Calibrator: 20 µl (Standard)
• Sample: 20 µl (Test)
• Distilled water: 20 µl (Blank)

Mix and allow to stand at room temperature (21–25°C) for 20 minutes or 5 minutes at 37°C. Read absorbance at 546 nm (530–570 nm) against distilled water. 

Calculations

Total Proteins (gm/dl) = (Aₜ – Aᵦ) / (A꜀ – Aᵦ) × Total Protein concentration provided on calibrator label 

Expected values

Total Proteins: 6.0 – 8.5 gm/dl

It is recommended that each laboratory should establish its own normal range representing its patient population. 

Performance 

• Linearity: 10 gm/dl 
• Comparison: r = 0.98; y = 1.4x + 0.6 
• Precision: within-run and run-to-run CV% values provided in IFU table (Low/High levels). 

Automated applications

VDx Total Protein liquid reagents can be used with (examples listed in IFU): Hitachi 700 series, RA 1000/2000/XT, Express 550 plus, Syncron CX4, Lisa 200, BTR 810/820/830, RA50, Erbachem-5 plus, etc. Application sheets for specific semi-automatic, batch and auto analysers are available on request. 

Typical input parameters: end point reaction; wavelength 546 nm; incubation 20 minutes / 5 minutes; calibrator concentration provided on label; temperature 21–25°C / 37°C; units gm/dl. 

Precautions (selected)

• For in-vitro diagnostics use only.
• Do not inhale the reagent or pipette by mouth.
• Do not freeze or expose reagents to higher temperature as it may affect performance.
• Before the assay bring reagents to room temperature.
• Avoid contamination of reagent during the assay process.
• Do not use reagent if it is hazy or cloudy. 

Quality control

For accuracy, it is necessary to run known serum controls with each assay. 

Frequently Asked Questions

What are protein reagents used for?

This reagent kit is used for in-vitro quantitative determination of total protein in serum and plasma. 

Which method is used in these protein reagents?

The kit uses the Biuret method (end-point), measured at 546 nm (530–570 nm). 

What is the measuring range (linearity) stated in the IFU?

Linearity is stated as 10 gm/dl. 

What are the expected total protein values?

The IFU lists 6.0–8.5 gm/dl as expected values and recommends each lab establish its own reference range.