Electrophoresis: 200 Tests R1 R2 R3 R4

ELECTROPHORESIS Hb SET.
SET COMPLETE FOR QUALITATIVE AND QUANTITATIVE DETERMINATION OF HAEMOGLOBINS VARIANTS

R1 1 x 50 ml., R2 1 x 250 ml., R3 1 x 100 ml., R4 1 x 25 ml., +
REF. SL-220-HB 200 Tests Strips in Cellulose Acetate supported 76 x 60 mm. 1 x 25 pcs.
Wicks 4 pcs. , Absorbents papers 17 x 59 mm. 1 x 8 pcs.
R1 1 x 50 ml., R2 1 x 250 ml., R3 1 x 100 ml., R4 1 x 25 ml.,
REF. SL-80-HB 80 Tests + Strips in Cellulose Acetate supported 76 x 60 mm. 1 x 10 pcs.
Wicks 4 pcs. , Absorbents papers 17 x 59 mm. 1 x 8 pcs.
Kits dedicated to the Instruments: ADALYA 24 - SELVET 24 - MINIPHOR 08 – and ADALYA 40
PRINCIPLE OF METHOD
The haemoglobins variants are separated electrophoretically on cellulose acetate plates in a specific buffer medium. After separation
the fractions are fixed, stained and destained by the reagents contained in the kit, in order to visualize the 2 haemoglobins variants:
HbA, HbA2 and in case the pathological variants HbF and HbS. The following optical-densitometric reading gives the percent value of
each fraction
REAGENTS
In vitro diagnostic medical device. IVD
The kit stored at 15-25 °C and protect from light, is stable up to expiry date shown on the package.
Components and concentration of final working mixture.
REAGENT R1 ( 10 x ) REAGENT R2 Destaining ( 20 x ) Xi

TRIS Buffer/ Glycine pH 9.2 Citric Acid 1 M/L R-37/38 , R-41
EDTA- Di Sodic Sodium phosphate bi-basic 1 M/L S-26 , S-36/37/39
Sodium Azide ( 0.02%)
REAGENT R3 Staining (ready to use)

Ponceau Red S 0.2 % (p.v.) R-35, R-36/38
Trichloroacetic Acid 3 % (p-v.) S-26 , S-24/25
REAGENT R4 Lysing solution (ready to use) Others materials supplied to the SET EL040 EL030
Phosphate Buffers pH 7.2 Cellulose Acetate Strips 76 x 60 mm. 1 x 25 pcs 1 x 10 pcs
Non ionic tensioactive 1% Wicks 1 x 4 pcs 1 x 4 pcs
Sodium Azide ( 0.02%) Absorbing papers 1 x 8 pcs 1 x 8 pcs
SAMPLES
TIPOLOGY : Red Cells ( Haemolysed ) starting by whole blood collected with EDTA
STABILITAY of SAMPLES ; 1 week stored at 2-8 °C.
MODALITY : For the taking of serum samples follow the indications container in NCCLS H11-A3.
MATERIAL REQUIRED FOR EXECUTION OF TEST BUT DON’T FURNISHED WITH THE KIT
Instruments : ADALYA 24 - SELVET 24 - MINIPHOR 08 and ADALYA 40 + usual equipment for Clinical Chemistry of
Laboratory.
Instructions : Use only mono-use material to avoid contaminations.
Use bi-distilled water stored in a suitable plastic container, for the dilution of Reagents R1 and R2.
PREPARATION OF WORKING SOLUTION
The REAGENT R1, Buffer, is 10 times concentrated. Dilute in ratio 1:10 with distilled water.
The dilute solution is stable up to the expiry date already reported on the label of concentrated solution, if stored at 2-8 °C .
The REAGENT R2, DESTAINING , is 20 times concentrated. Dilute in ratio 1:20 with distilled water. To easy the dilution is advisable
to extend at 2 litres, with distilled water, the whole content of 100 ml vial. The dilute solution is stable up to the expiry date already
reported on the label of concentrated solution.
PREPARATION of SAMPLES
Preparation of HEMOLYSATE
1)In a conical bottom tube, pipette a volume of red blood cells to which 9 volumes of physiological saline
should be added. Gently invert several times and centrifuge at 2000 rpm for 5 minutes.
2) Remove the supernatant liquid, add 9 volumes of physiological saline again, gently invert several times
and centrifuge at 2000 rpm for 5 minutes.
3) Repeat the operation, as described in point 2, until the supernatant is clear and colorless.
4) Add 3 parts of Lisante solution (Reag.R4) to one part of washed erythrocytes. Gently agitate the tube and
wait for 5 minutes. Use as a sample the cell red cell extract solution (hemolysate).
________________________________________________________________________________________________
SELEO s.r.l. Headquarter : I Traversa Bugnano s.c. 81030 – Orta di Atella – (CE) ITALY tel +39( 0)815022877 r.a. fax+ 39(0)81502287702
P.IVA e C.F. IT 08850061212 web site : www.seleo.it
5) Dispense 25/30 microliters of EMULISATE, thus prepared, into each well of the sample holder base,
ensuring homogeneous distribution and avoiding the formation of bubbles
6) Warning: to obtain a perfect standardization of the sample, measure the concentration of Hb present in
the hemolysate, using a Hematology device, and possibly dilute with distilled water, to obtain a final
concentration of Hb equal to 4 g / dL.
PROCEDURE dedicated for the Instruments : ADALYA 24 - SELVET 24 - MINIPHOR 08 and ADALYA 40
Insert the vial of REAGENT R3, Staining (Ready to use), in reagents place of instruments ADALYA 40 or SELVET 24, assuring the
insert of loading tube labelled with the “ STAIN “ word. The vial content is useful for the right coloration of 25 cellulose acetate
membranes.
In the electrophoretic chamber of the instruments, dispense up to the shown level ( ca. 150 ml. ) the solution diluted 1:10 of Buffer
( Reag. R1), and insert the 2 wicks in the apposite lodges.
Buffer is stable up to the utilize of 25 membranes of cellulose acetate (if stored at 2-8 °C). Is advisable, at the end work, to extract from
the instrument the Electrophoretic chamber and to keep
Replace the solution after 25 migrations or in case of any bacterial growth or moulds.
Drop the remaining buffer solution in the bottle placed in the system reagent box marked “BUFFER SOLUTION”; this reagent will allow
the automatic soaking of 25 electrophoretic plates.
In the Electrophoresis chamber of the instrument, dispense up to the shown level (ca. 150 ml ) the solution diluted in ratio 1:10 of
Reagent R1 buffer. Change the solution after 25 acetate plates or in case of moulds formation. Insert in the apposite lodges of
Electrophoresis chamber, the wicks (included in the kit).
Dilute the REAGENT R2 , Destaining, with distilled water in ratio 1 :20 , 1 vial of 250 ml, extend at 5 litres in polyethylene tank
(supplied). Insert in the tank, the loading tube for the destaining. Be sure that the tank level is never lower than 500 ml during the use.
Have therefore a tank containing five litres of distilled water for the washing of instrument “ CLEAN “, and a tank (empty) for the
discharge labelled as “ WASTE “.
Mount the cellulose acetate membrane, by the apposite frame strips-holder, avoiding to touch the surface. Insert the membrane in the
strips-holder rack, starting from the position 1. Insert the absorbing papers into the lodges.
FOR MORE DETAILS SEE THE MANUAL OF USE SPECIFIC OF THE INSTRUMENT
OPERATIONS / PARAMETERS Migration Voltage ⇒ 220 V Optical Compensation ⇒ 25
SOAKING ⇒ 180 seconds DRYING 1 ⇒ 90 seconds MIGRATION ⇒ 1500 seconds

COLORATION ⇒ 180 seconds DESTAINING 1 ⇒ 360 seconds DESTAINING 2 ⇒ 480 seconds
DRYING 2 ⇒ 30 seconds DEPOSITION time ⇒ 25 seconds
PERFORMANCE OF SYSTEM
Newborn : Hb A1 = 10 – 40 % , Hb A2 = < 0.2% , Hb f = 60 -90%

Adult : Hb A1 = 96.9 – 98.9 % , Hb A2 = 1.1 – 3.1% , Hb f = < 1% ( up to 5 years : 0 – 5%, beyond : 0 – 2%)
Hb S : Falciform anemia < or missing HbA1 ( with possible increasing of HbF ).

Talassemia > Hb A2 e < Hb A1
Talassemia MAJOR : < Hb A1 > Hb f ( 70 – 90 %) Talassemia MINOR : slight increasing Hb f ( up to 10% ) > Hb A2 ( 2,3 times)
PRECAUTION OF USE
The reagents contain harmful substances as indicated on the label in accordance with EEC Directives 67/548 / EEC and 88/379 / EEC
on the Classification, Packing and Labeling of Hazardous Substances. It is recommended to follow the rules of good laboratory practice
in the use of reagents. Handle with caution, avoid swallowing, avoid contact with skin, eyes and mucous membranes.
See the Safety Sheet specific for the Reagent R1 and R2.
REGULAMENTATION INFORMATION D. Lgs 14/3/2003 n°65, D.Lgs.03/02/1997 n°57 (Classification and Labelling )
Phrases R : R-37/38 Irritating to respiratory system and skin, R-41 Risk of serious damage to eyes R-35 Cause severe burns
Phrases S: S-24/25 Avoid the contact with skin and eyes
S-26 In case of contact with eyes, wash immediately with plenty of water and seek medical advice.
S-36/37/39 Wear suitable protective clothing , gloves and eye/face protection
WASTE DISPOSAL
Expired or used products must be disposal as special waste of laboratory.
SIMBOLOGY
In vitro diagnostic medical device
Manufacturer Dimension/number test
CE mark (product conforming at Dir. 98/79/CE)
Stored temperature To follow instructions
Xi Irritant C. Corrosive ELECTROPHORESIS

HB SET
________________________________________________________________________________________________
SELEO s.r.l. Headquarter : I Traversa Bugnano s.c. 81030 – Orta di Atella – (CE) ITALY tel +39( 0)815022877 r.a. fax+ 39(0)81502287702
P.IVA e C.F. IT 08850061212 web site : www.seleo.it

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ELECTROPHORESIS Hb SET.

SET COMPLETE FOR QUALITATIVE AND QUANTITATIVE DETERMINATION OF HAEMOGLOBINS VARIANTS

Kits dedicated to the Instruments: ADALYA 24 - SELVET 24 - MINIPHOR 08 – and ADALYA 40

REAGENT R1 ( 10 x ) REAGENT R2 Destaining ( 20 x ) Xi

REAGENT R3 Staining (ready to use)

SOAKING ⇒ 180 seconds DRYING 1 ⇒ 90 seconds MIGRATION ⇒ 1500 seconds

Newborn : Hb A1 = 10 – 40 % , Hb A2 = < 0.2% , Hb f = 60 -90%

Hb S : Falciform anemia < or missing HbA1 ( with possible increasing of HbF ).

In vitro diagnostic medical device

Manufacturer Dimension/number test

CE mark (product conforming at Dir. 98/79/CE)

Stored temperature To follow instructions

Xi Irritant C. Corrosive ELECTROPHORESIS

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