Basic Concepts in Clinical Biochemistry-A Practical Guide.7z

(Chris Devlin) #1

  1. Pipette out 25, 50, 75, 100, 125, 150, 175, and 200μl of 4 mg % calcium standard
    into the tubes labelled as S 1 to S 6. Add 50μl undiluted serum and urine samples
    into tubes Ts and TU.

  2. Then make thefinal volume of all the tubes 200μl by adding distilled water. Add
    2 ml cresolphthalein complexone reagent to all the tubes and mix well.

  3. Then add 2.5 ml diethylamine buffer to all the tubes, mix well, and measure
    absorbance at 575 nm.


Reagents Blank S 1 S 2 S 3 S 4 S 5 S 6 S 7 S 8 TS TU
Calcium standard
(μl)


  • 25 50 75 100 125 150 175 200 ––


Calcium amount
(μg)


  • 123456 78––


Serum/urine (μl) ––––––– 50 50
Distilled water
(μl)

200 175 150 125 100 75 50 25 – 150 150

Cresolphthalein
complexone
reagent (ml)

2 222222 2222

Diethylamine
buffer (ml)

2.5 2.5 2.5 2.5 2.5 2.5 2.5 2.5 2.5 2.5 2.5

Mix well and measure absorbance at 575 nm

21.7 Calculations


Plot a graph by taking calcium amount at x-axis vs absorbance at y-axis, and
extrapolate calcium concentration of given serum/urine sample from the graph.
The calcium levels in serum and urine can also be calculated by using equation:


In serum¼
ODof testamount of standardðÞμg 100
ODof standardvolume of sample 1000
¼xmg=dl

In urine¼

OD of testamount of standardðÞμg 1000
OD of standardvolume of sample 1000
¼xmg=L

21.8 Clinical Significance...............................


The normal value of calcium in serum is 9–11 mg/dl and 100–300 mg/24 h urine.
Extracellular calcium provides calcium ions for maintenance of intracellular cal-
cium, bone mineralization, blood coagulation, and maintenance of plasma


21.8 Clinical Significance 87

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