Veterinary Clinical Biochemistry and Laboratory Diagnosis

PROCEDURE

Estimation of serum creatinine - Alkaline Picrate Method

Sample

• Serum /Plasma

Reagents:[reagents in the kit vary from manufacturer to

                  manufacturer so the procedure in  the kit used may be followed.]

• Picric acid reagent
• Sodium hydroxide 0.75 N
• Creatinine Standard (150 mg%)

Preparation of working Standard

• Dilute 0.1 ml of urea Reagent to 10 ml with distilled water.

Procedure (Agappe Diagnostics Kit)  

• Deproteinization of Serum
  • Pipette into a clean dry test tube (T)-serum 0.5ml, distilled water-0.5 ml and picric acid reagent-3.0 ml .
  • Mix well and centrifuge at 3000 rpm for 10 minutes to obtain a clear supernatant.

Colour Development

• Pipette into a clean dry test tubes labeled blank (B), Standard (S) and test(T) as follows

S.No	Reagents	(B) (ml)	(S) (ml)	(T) (ml)
1.	Standard creatinine	-	0.5	-
2.	Supernatant from above step	-	-	2.0
3.	Distilled water	0.5	-	-
4.	Picric acid reagent	1.5	1.5	-
5.	Sodium hydroxide 0.75 N	0.5	0.5	0.5

• Mix well and allow the reaction mixture to stand at room temperature for exactly 20 minutes.
• Measure the absorbance of the standard (S) and test (T), blank (B) against Distilled water at 520 nm or with a filter in a photometric colorimeter.

Calculation

Results

• The creatinine content of the given serum is -----------mg/dl