Summary

Calcium, in the body, is found mainly in the bones

(approximately 99%). In serum calcium exists equally in

a free ionised form and in a bound form (with albumin).

Hence, a decrease in albumin causes lower calcium levels

and vice versa. The levels of calcium in serum depend on

the parathyroid hormone. Increased calcium levels are

found in bone tumors, hyperparathyroidism. Decreased

levels are found in hypoparathyroidism, renal failure,

rickets, vitamin D deficiency and pancreatitis.

Principle

Calcium in an alkaline medium combines with

O-Cresolphthalein complexone to form a purple colored

complex. Intensity of the color formed is directly proportional

to the amount of calcium present in the sample.

Calcium + OCPC Purple colored complex

Normal Reference Values

Serum/plasma : 8.7–11.0 mg/dL (general)

It is recommended that each laboratory establish its

own normal range representing its patient population.

Contents 2 × 35 mL 2 × 75 mL

L1: Buffer reagent 35 mL 75 mL

L2: Color reagent 35 mL 75 mL

S: Calcium standard (10 mg/dL) 5 mL 5 mL

Storage/Stability

Contents are stable at 2–8°C till the expiry mentioned on

the labels.

Reagent Preparation

Reagents are ready to use. Protect from bright light.

Working reagent: For convenience a single working

reagent may be prepared by mixing equal parts of the

buffer reagent and color reagent. This combined reagent

is stable for 7 days at 2–8°C.

Sample Material

Serum/heparinized plasma. Calcium is reported to be

stable in serum for 7 days at 2–8°C.

Contd...

494 Concise Book of Medical Laboratory Technology: Methods and Interpretations Procedure

Wavelength/filter : 570 nm (Hg 578 nm)/yellow

Temperature : RT

Light path : 1 cm

Pipette into clean dry test tubes labeled as blank (B),

standard (S), and test (T):

Addition

Sequence

B

(mL)

S

(mL)

T

(mL)

Buffer reagent ( L1 ) 0.5 0.5 0.5

Color reagent ( L2 ) 0.5 0.5 0.5

Distilled water 0.02 - -

Calcium standard (s) - 0.02

Sample - - 0.02

Mix well and incubate at RT (25°C) for 5 minutes.

Measure the absorbance of the standard (Abs S), and test

sample (Abs T) against the blank, within 60 minutes.

Calculations

 Abs T

Calcium in mg/dL = ________ × 10 Abs S

Linearity

This procedure is linear upto 18 mg/dL. If values exceed

this limit, dilute the sample with distilled water and repeat

the assay. Calculate the value using the proper dilution.

Note

As calcium is a very widely distributed ion, care should

be taken to avoid any contamination. All glassware being

used for the test should first be rinsed with 1% or 0.1 N

HCI and then with good quality deionized water before

use.

It is suggested that after the rinsing of the tubes with

HCI the reagent be pipetted in their respective tubes

and the tubes be rinsed with the reagent. The reagent

then should be pooled together in the ‘blank’ tube and

repipetted out into the ‘standard’ and ‘test’ test tubes.

This will ensure that any remaining contamination will be

carried over equally in all the tubes. For flow cell cuvettes,

it is suggested that some reagent be aspirated before the

blank to take away any contamination in the flow through

tubing or cuvette which may cause a higher than the actual

blank of the reagent.

Chelating agents such as EDTA, present even in

traces, prevent the formation of the color complex, hence

necessary care should be taken during the assay.

System Parameters

Reaction : End point Interval :

Wavelength : 570 nm Sample volume : 0.02 mL

Zero setting :  Reagent

blank

Reagent volume : 1.00 mL

Incubation

temperature

: RT Standard : 10 mg/dL

Incubated time : 5 min Factor :

Delay time : — React slope : Increasing

Read time : — Linearity : 18 mg/dL

No. of read : — Units : mg/dL

Calcium (Arsenazo III Method)

(Courtesy: Tulip Group of Companies)

For the determination of calcium in serum or plasma (for

in vitro diagnostic use only).

Summary

Calcium, in the body, is found mainly in the bones

(approximately 99%). In serum calcium exists equally in

a free ionized form and in a bound form (with albumin).

Hence, a decrease in albumin causes lower calcium levels

and vice versa. The levels of calcium in serum depend on

the parathyroid hormone. Increased calcium levels are

found in bone tumors, hyperparathyroidism. Decreased

levels are found in hypoparathyroidism, renal failure,

rickets, vitamin D deficiency and pancreatitis.

Principle

Calcium combines specifically with arsenazo III at a

neutral pH to form a blue purple colored complex.

Intensity of the color formed is directly proportional to the

amount of calcium present in the sample.

Calcium + Arsenazo III Blue Purple

 colored complex

Normal Reference Values

Serum/plasma : 8.7–11.0 mg/dL

It is recommended that each laboratory establish its

own normal range representing its patient population.

Contents 75 mL 3 × 75 mL

L1: Calcium reagent 75 mL 3 × 75 mL

S: Calcium standard (10 mg/dL) 5 mL 5 mL

Storage/Stability

Contents are stable at 2–8°C till the expiry mentioned on

the labels