VLD 421: Veterinary Clinical Biochemistry and Laboratory Diagnosis-II (0+1)

ESTIMATION OF TRIGLYCERIDES

Principle

• Triglycerides are hydrolysed by the lipase to glycerol and free fatty acids.
• Glycerol is phosphorylated by ATP in the presence of glycerolkinase to glycerol -3-P, which is oxidised by Gly-3-P oxidase (GPO) producing H₂O₂.
• H₂O₂ so formed reacts with 4-amino -antipyrine and 3,5 -dicholoro-2-hydroxy benzene sulfonic acid in the presence of peroxidase (POD) to produce a red quinoneimine dye.
• The intensity of coloour developed is proportioinal to the triglyceride concentration.

Procedure

Specimen

• Serum/Plasma

Reagents

• Enzyme reagent
• Buffer solution
• Triglycerides standard (200mg/dl)

Procedure (Agappe Diagnostic Kit)

• Pipette into clean, dry acid -washed test tubes labeled as blank(B), stanard (S) and test (T) as follows:

S.No	Reagents	B (ml)	S (ml)	T (ml)	OD at 510
1	Working enzyme reagent	0.5	0.5	0.5
2	Triglyceride standard	-	0.01	-
3	Serum	-	-	0.01
Mix well and incubate at 37 c for 10 minutes
4	Distilled water

• Measure the absorbance of standard (S) and test (T) against blank (B) at 510nm or with a green filter in a photoeletric colorimeter within 30 minutes .
  

Calculation

Serum Triglycerides (mg/dl) =    (O.D of test / O.D of std ) x 200

Result

• The content of triglycerides in the given serum = mg/dl

Clinical Significance:

• Levels of both cholestrol and triglycerides in blood have been idendified as risk factors related to atherosclerotic disease . the levels of cholestrol and triglycerides can vary independantly and hence evaluation of hyperlipidemia includes determination of both levels.
• Elevated levels of glycerides are found in atherosclerotic disease,Diabetics mellitus, biliary obstruction and other metabolic disorders associated with endocrine distrubances.