Citric Acid Test the total volume of the semen sample or seminal plasma.

PHOTOMETRIC TEST FOR QUANTIFYING CITRIC ACID IN HUMAN TEST - RETEST RELIABILITY
SEMINAL PLASMA
4 different samples where each tested 10 times:
Document reference: FP09 I37 R01 A.1
Average (mg/mL) 10.69 10.97 10.65 11.91
Update: 21/01/2004
S.D. 0.521 0.581 0.796 0.648
In Vitro Diagnostic V.C. % 4.87 5.29 7.47 5.44
Reagent for Professional Use only.
5 samples where spiked with 0, 1, 2 and 3 mg citric acid
GENERAL INFORMATION and then tested with the Citric Acid Test. The testresults
of the unspiked sample where subtracted from results of
The secretory products of the prostate gland form about one-
third of the seminal fluid in which spermatozoa are sus- the spiked sample and then set out against the standard:
pended. The level of Citric Acid in semen gives a reliable
Spike 0 1 2 3
measure of prostate gland secretion (Comhaire, 1986).
Observed 0 1.2 1.9 3.4
TEST PRINCIPLE Recovery - 120% 95% 115%
The Citric Acid test works in two steps:
STORAGE
1. Spermatozoa and particles are removed by isopropa-
Store reagents between 2°C and 25°C. Keep from light.
nol
Product can be used for 12 months after date of produc-
2. After centrifugation, ferric chloride is added to the
tion.
supernatant. The Fe3+-ions and citrate form a complex
that turns the solution to a yellow colour. The intensity of WARNINGS AND PRECAUTIONS
the colour is directly related to the amount of citrate and
Reagent 1: Harmfull if swallowed. Irritating to skin. Risk of
can be measured in a photometer or plate reader. serious damage to eyes.
MATERIAL INCLUDED IN THE KIT Reagent 2/3: Highly flammable. Irritating to eyes. Vapours
may cause drawsiness or dizziness.
Reagent 1 - 20mL FeCl3 solution (Foams easily: Do not
shake) All human, organic material should be considered poten-
Reagent 2 - 10mL Iso-propanol tially infectious.
Reagent 3 - 2mL Citric Acid standard (4mg/mL) Handle all specimens as if capable of transmitting HIV or
hepatitis. Always wear protective clothing when handling
MATERIAL NOT INCLUDED IN THE KIT specimens and reagent (gloves, lab vest, eye/face protec-
Plate reader / Photometer (405nm filter) tion).
Pipettes and tips
Centrifuge tubes CALCULATION OF G-FORCES
Titre plate The g-force of your centrifuge can be calculated using this
Centrifuge (+1500g) formula:
Small reagent tubes or Eppendorf tubes
g = 1.118 x r x rpm² or rpm = Square root {g / (1.118 x r)}
METHOD
r = radius of centrifuge in mm
1. Mix 100µL of Reagent 2 with 100µL of semen or rpm = rotations per minute / 1000
semen plasma and mix
e.g.1
2. Standard: mix 100µL of standard solution with 100µL
r = 100 mm rpm = 3000 rotations per minute
of Reagent 2
3. Centrifuge for 10 minutes at 1500g’s or more g = 1.118 x 100 x 9 = 1006g
4. Pipette 25µL of supernatant into an empty well. e.g.2
Note: Remove supernatant very carefully in order not to r = 100 mm g = 1200g
aspire any of the sediment. In case the supernatant rpm = SQR {1200 / (1.118 x 100)} = 3.28
is turbid, centrifuge again at a higher speed for = 3280 rotations per minute
another 10 minutes.
BIBLIOGRAPHY
5. Add 200µL of Reagent 1 to the well, slowly
6. Read the results of the samples and the standard at F.H. Comhaire; Male Infertility. Clinical investigation, cause
evaluation and treatment - 1st Edition; Chapman & Hall;
405nm
1986.
INTERPRETATION
The measured value (OD) for the sample is devided by the Citric Acid Test is manufactured by :
OD from the standard and multiplied by the concentration of FertiPro N.V.
the standard (4mg/mL). Industriepark Noord 32
ODSample 8730 Beernem - Belgium
Citric Acid mg/mL = x 4mg/mL E-mail: info@fertipro.com
ODStandard URL: http://www.fertipro.com

To obtain total Citric Acid amounts, multiply the result with