Creatinine

Creatinine

Kinetic colorimetric method
FIXED TIME

PRINCIPLE

This procedure is based upon a modification of the original picrate reaction (Jaffe). Creatinine under alkaline conditions reacts with picrate ions forming a reddish complex. The formation rate of  the complex measured through the increase of absorbance in a prefixed interval of time is proportional to the concentration of creatinine in the sample.

REAGENT COMPOSITION

4 x 100 ml
R1 Picric acid: Picric acid 25 mmol/L
R2 Alkaline buffer: Phosphate buffer 300 mmol/L pH 12.7,
SDS 2.0 g/L (w/v). Xi R:36/37/38
CAL Creatinine standard: Creatinine 2 mg/dL (177 umol/L).
Organic matrix based primary standard. Concentration
value is traceable to Standard Reference Material 914a.

STORAGE AND STABILITY
Store at 2-30ºC

REAGENT PREPARATION
Working reagent. Mix 1 volume of R1 + 1 volume of R2. Stable for
1 week at room temperature, stored tightly closed and protected
from light.
SAMPLES
Serum or heparinized plasma, and urine (see Notes).
Creatinine in serum or plasma is stable up to 24 hours at 2-8ºC.
Freeze for longer storage.
Creatinine from random samples of urine is stable for 4 days at 2-
8ºC. Freeze for longer storage. The 24-hour urine samples for the
Clearance Test should be collected on a preservative (fluoridethymol) and immediatelly refrigerated.

MATERIALS REQUIRED

• Photometer or colorimeter with a thermostatic cell compartment, able of reading at 510 ± 10 nm.
• Constant temperature incubator set at 37ºC.
• Stopwatch, strip-chart recorder or printer.
• Cuvettes with 1-cm pathlength.
• Pipettes to measure reagent and samples.

Creatinine reagent per box