Made in France Latest revision : www.biolabo.

fr Revision : 23/04/2013


STONE ANALYSIS SET
Chemi cal qual i t at i ve met hod

Reagent for qualitative determination of main
individual components of urinary stones (urinary calculi).

REF 92315
REF 92330
100 tests
40 tests

TECHNICAL SUPPORT AND ORDERS
Tel : (33) 03 23 25 15 50
Fax : (33) 03 23 256 256
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IVD IN VITRO DIAGNOSTIC USE

CLINICAL SIGNIFICANCE (1) (2)
The presence of calculus in urines is due to the conglomeration and
the crystallization of matter in supersaturation. There can be various
reasons for this imbalance: nutritional, metabolical, genetical,
anatomical, iatrogenical, neurological or infectious reasons. Several
ones can occur, at the same time or successively, to the formation and
the growth of the calculus.
The more frequently met chemical components are (in descending
frequency order) : calcium oxalates, calcium phosphate and
magnesium ammonium phosphates, uric acids and urates, various
proteins, cystine.
The qualitative analysis of a certain components (among the most
frequent ones) of urinary calculus represents a first approach to the
etiological diagnosis of urinary lithiasis and the direction of the patients
therapeutical follow-up. It is absolutely necessary to perform the test
together with morphological analysis (low power stereo microscope,
light microscopy) and/or with molecular and crystallin identification (X-
ray diffraction, infrared spectophotometry). Because of its nature, the
calculus has to be the subject of a precise and reliable analysis.
Coupled and rationally used optical and chemical methods can, in
most cases, give sufficient information about the composition and the
etiology of the calculus.
PRINCIPLE (4) (5)
This method is used for identification of main mineral components and
one organical component (cystine) of urinary calculi by easy chemical
tests.
REAGENTS COMPOSITION
vial R1 :
Hydrochloric Acid (HCl 1,65 M)
Corrosive, R34, S36/37/39
vial R2
Sodium Hydroxyde (NaOH 6,25 M)
Corrosive, R35, S36/37/39
vial R3
1
er
Reagent for Cystine determination
(NaOH, Sodium Cyanide)
Corrosive, Toxic, Harmful for environment
R34, R23/24/25, R31-52/53, S36/37/39, S46
vial R4
2
nd
REAGENT FOR CYSTINE DETERMINATION
(Sodium nitroprussiate)
Xn: Harmful, R22, S21/26/37
vial R5
Reagent for Phosphates determination
(Sulfuric Acid, Ammonium Molybdate, Ferric Sulfate)
Corrosive, R34, S36/37/39
vial R6
Reaget for Magnesium determination
(NaOH, paranitrophenylazoresorcinol)
vial R7
Reagent for Calcium determination
(KOH, calcein)
Xi: Irritating, R34, S36/37/39
vial R8
Reagent for Ammonia determination
(Potassium Iodide, Mercuric Iodide)
Corrosive, R36/38, S36/37/39
vial R9
Reagent for Uric Acid determination
(Acetic Acid, neocuprone, Copper Sulfate)
Xi: Irritating, R36/38, S36/37/39
vial R10
Reagent for Oxalate determination
(Manganese dioxide)
Xn: Harmful, R20/22, S36/37/39, S22-26-281
SPATULA
For use with R10 reagent only. Store and wipe away dust after use
SAFETY CAUTIONS
See also REAGENTS COMPOSITON and respect the symbol of
security indicated on the label of the vial.
Refer to Material Safety Data Sheet to know the significance of R and
S Phrases.
BIOLABO reagents are designated for professional, in vitro diagnostic
use.
Verify the integrity of the contents before use.
Use adequate protections (overall, gloves, glasses).
Do not pipette by mouth.
In case of contact with skin and eyes, thoroughly wash affected
areas with plenty of water and seek medical advice.
Waste disposal : Respect legislation in force in the country.
All specimens should be handled as potentially infectious, in
accordance with good laboratory practices using appropriate
precautions. Respect legislation in force in the country.
REAGENTS PREPARATION
Reagents are ready for use.
STABILITY AND STORAGE
Store away from light, well cap in the original vial at 18-25C.
When free from contamination, stored and used as described in the
insert, reagents are stable until expiry date stated on the label of the
kit.
SPECIMEN COLLECTION AND HANDLING
Morphological studies should be made with a whole calculus cleared
out of possible impurities. Qualitative analysis and flame test should be
realised on pulverised calculus using clean pestle and mortar to obtain
finely ground powder.
INTERFERENCES (4)
Cobalt and nickel do not interfere with the determination of the
magnesium because of their weak concentration in the organism.
Carbonate testing : if there has been effervescence during the
addition of R1 revealing the presence of carbonate, add R1 drop by
drop until the end of the gas evolution. Then, shake vigorously
("Vortex") for at least one minute so as to get rid of all the carbon
dioxyde (or failing that, bring to the boil a few seconds in a Pyrex
tube and bring back at room temperature). This process is necessary
to avoid a false-positive result during the determination of oxalate.
MATERIAL REQUIRED BUT NOT PROVIDED
1. Low power stereo microscope
2. Clean porcelain pestle and mortar
3. Metallic seeding loop
4. mg scale (weighing of the calculus powder)
5. Pipette to dispense 1 drop (approx. 50L) of reagents and mixture
M1, M2.
6. Tubes, or glass/ceramic cored plate with a white bottom

BIOLABO REAGENTS
www. b i o l a b o . f r
MANUFACTURER:
BIOLABO SA,
02160, Maizy, France

Made in France Latest revision : www.biolabo.fr Revision : 23/04/2013
QUALITY CONTROL
When calculi show an unusual morphology or lead to negative or
incoherent results during the chemical analysis, one should carry out
more appropriate analysis that could highlight a particular composition
or etiology.
It is recommended that each laboratory establishes its own
investigation procedure by technics adapted to the diversity not only of
the structure, but also of the molecular composition of the calculus
studied.

Specificity : Each reaction is specific to the tested component. For
oxalate, see also INTERFERENCES.
PERFORMANCE CHARACTERISTICS
Detection limits :
Carbonate : 1 mg of Calcium carbonate
Cystine : 1 mg of L-Cystine
Phosphate : 1 mg of Calcium phosphate
Ammonium : 1 mg of Ammonium magnesium Phosphate
Magnesium: 3 mg of Ammonium magnesium Phosphate
Calcium: 0.1 mg of calcium (phosphate, carbonate, oxalate)
Uric acid : 0.1 mg of uric acid
Oxalate : 2.5 mg of Calcium Oxalate
Sensitivity : Reagents are very sensitive and so allow the detection of
the different components even as trace

PROCEDURE (1)

Morphological analysis of calculus should be made before pulverizing the calculus for chemical test.
Flame test : Immerse a metallic seeding loop into distilled water, then into the calculus powder. Bring to the flame. The carbonization or the
disappearance of the powder is the sign that the components are mostly of organic origin. The absence of carbonization shows that the
calculus is of mineral origin.
Qualitative chemical tests : (see table below)
Step 1 : Weigh about 50 mg of the calculus powder, transfer into a test tube and add 10 drops of reagent R1. An effervescence shows the
presence of carbonate (see INTERFERENCES). In this case, shake vigorously for 1 minute. The remaining mixture is called M1 in the
table below.
Preparation of the mixture M2 : Mix 50 l of M1 and 5 ml of distilled water. Mix well and use only for the determination of calcium (Step 5).
According to the step below, dispense 1 drop (approx. 50 L) of the mixture M1 or M2 in each cavity of a ceramic cored plate or in tubes
and carry out the next tests (Step 2 to 8) :

Step 1 Step 2 Step 3 Step 4 Step 5 Step 6 Step 7 Step 8
CARBONATE CYSTINE PHOSPHATE MAGNSIUM CALCIUM AMMONIUM URIC ACID OXALATE

CALCULUS
POWDER
50 mg
+
R1
10 drops
(500 L)

= M1

1 DROP
M1
1 DROP
M1
1 DROP
M1
1 DROP
M2
1 DROP
M1
1 DROP
M1
REMAINING
MIXTURE
M1
+
R3
1 DROP
+
R5
2 DROPS
(100 L)
+
R6
1 DROP
+
R2
1 DROP
+
R2
1 DROP
+
R2
1 DROP
+
R10
THE TIP OF A
SPATULA
MIX,
LET STAND
FOR 5 MIN.
MIX,
LET STAND
FOR 5 MIN.
+
R2
5 DROPS
(250 L)
+
R7
2 DROPS
(100 L)
+
R8
1 DROP
+
R9
1 DROP
Approximately.
60 mg
+ R4
1 DROP






MIX


MIX


MIX


MIX

WAIT FOR
FEW SECONDS

POSITIVE
RESULT
EFFERVESCENCE
NOTE 2
RED COLOUR BLUE COLOUR BLUE
PRECIPITATE
YELLOW
COLOUR
ORANGE-BROWN
PRECIPITATE
YELLOW/ORANGE
COLOUR
EFFERVESCENCE
NOTE 2
NEGATIVE
RESULT
ABSENCE OF
EFFERVESCENCE
YELLOW COLOUR NO CHANGE IN
COLOUR
ABSENCE OF
PRECIPITATE
PURPLE COLOUR
ORANGE
COLOUR
YELLOW COLOUR NO CHANGE IN
COLOUR
ABSENCE OF
EFFERVESCENCE

Notes :
1) Negative control : for steps 2 to 7, it can be useful to make a negative control by replacing the mixture M1 or M2 (according to the test) by
demineralised water so as to better differentiate the change in colour.
2) Effervescence : visible and audible
REFERENCES
(1) Les calculs urinaires : M. DAUDON, le Biotechnologiste, n4, (02/1994), p.8 11.
(2) Comment analyser un calcul et comment interprter un rsultat : M. DAUDON, lEurobiologiste (1993), 27, n203, p.35-46
(3) Revue critique des mthodes danalyse des calculs urinaires M. DAUDON et R. J. REVEILLAUD., Actualits nephrologiques de lhpital Necker, Flammarion mdecine
sciences, d. Paris,(1985) p. 203-224
(4) Routine Analysis of urinary calculi : Rapid simple method using spot tests, J. H. WINER et MATICE M. R., J. Lab. Clin. Med. (1943), 28, p.898-904
(5) P-nitrobenzenazoresorcinol solution ; use in test reagent for Magnesium : WELCHER F., Chemical solutions (1966)p.244
(6) Practical Value of analysis of urinary calculi, WINER. J. H., J.A.M.A. (1959), Vol.169, n15, p.1715-1718
IVD REF LOT
Manufacturer Use by In vitro diagnostic Temperature limitation Catalogue number See insert Batch number Store away from light sufficient for dilute with