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Total Protein Test Procedure

The total protein test measures the total amount albumin and globulin in your body. It’s used as part of your routine health checkup. It may also be used if you have unexpected weight loss, fatigue, or the symptoms of a kidney or liver disease.

Total Protein Test Procedure

SGPT Also Known as : TP, Total Protein, T. Protein

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Test Panel: Total Bilirubin, Conjagated Bilirubin, Unconjugated Bilirubin, ALT, AST, ALP, Total Protein, Albumin, Globulin, A/G ratio, GGT,

Biuret Method

Photometric test according to biuret method

Principle :

Proteins form a violet blue color complex with copper ions in alkaline solution. The absorbance of the color is directly proportional to the concentration.

Kit Contents:

  • R 1 :
    • Sodium hydroxide —– 100 mmol/L
    • Potassium sodium tartrate ———— 17 mmol/L
  • R 2 :
    • Sodium hydroxide —– 500 mmol/L
    • Potassium sodium tartrate —- 80 mmol/L
    • Potassium iodide ———— 75 mmol/L
    • Copper sulphate ———– 30 mmol/L
  • Standard: ——- 5.0 g/dL

Test Requirements :

  • Test Tubes
  • Micropipettes
  • Tips
  • Bio-Chemistry analyzer
  • SGPT Test Kit
  • Calibrator

Reagent Preparation and Stability :

Sample Starter:
4 Part of R1 (20 ml)
1 Part of R2 (5 ml)
and Mix well
Avoid direct exposure to light. Stability of Sample starter reagent: 1 Year at 2 – 25 °C.

Specimen collection and handling:

  1. Non-hemolyzed serum, heparinized or EDTA plasma is recommended.
  2. Stability: 10 days at 2 – 8° C.

Analyzer Parameter Required :

Reaction Type Photometric
Wavelength 540 nm (534nm – 565nm)
Cuvette Temp 37°C, 20 – 25°C
Zero Setting Against reagent blank
Light Path 1 cm

Test Procedure:

Pipette into clean dry test tube labeled as Sample (S), Calibrator Label (C)m Blank as (B) and QC Label (QC) or as your required:

Metho 1

Specimin(B)(S)(C) / (QC)
R11.0 ml (1000 µL ) 1.0 ml (1000 µL ) 1.0 ml (1000 µL )
Sample 20 µL
Calibrator or QC20 µL
Dist. water 20 µL
Mix, read absorbance A1 after 1 – 5 min. at 20 – 25 °C/ 37 °C, then add:
R2 250 µL 250 µL 250 µL
Mix, incubate for 5 min. at 20 – 25°C/37°C and read absorbance A2 within 60 min.

∆A = (A2 – A1) sample or standard

Method 2 ( Sample Starter)

Specimin(B)(S)(C) / (QC)
Sample Starter Reagent 1.0 ml (1000 µL ) 1.0 ml (1000 µL ) 1.0 ml (1000 µL )
Sample 20 µL
Calibrator or QC20 µL
Dist. water 20 µL
Mix, incubate for 5 min. at 20 – 25°C/37°C and read absorbance against the reagent blank within 60 min.
∆A = A Sample/Standard

CALCULATION : With standard or calibrator

Total Protein g/dL = ∆A Sample / ∆A Std/Cal x Conc std/cal g/dL

NOTE: Samples having a very low or high activity show a very low initial absorbance as most of the NADH is consumed prior to the start of the measurement. If this is suspected then dilute the sample and repeat the assay

LINEARITY :

The procedure is linearity is lessthen 0.06 and upto 15.0 g/dl and depends on KIT.
if the activity exceeds this limits dilute the sample with normal saline (NaCl 0.9 %) and multiply result by dilution factor.

QUALITY CONTROL :

For accuracy it is necessary to run known controls with every assay.

Normal Value :

Serum : 6.4 to 8.3 g/dL.
Note: Each Laboratory or Each Company Kits should establish it’s own normal range.

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