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C Protocol bij de Europese Overeenkomst betreffende de uitwisseling van geneesmiddelen van menselijke oorsprong, zoals gewijzigd

Biological test: limits and methods

(See Item II, 3 of Annex above):

(a) Limit:

A salt solution equivalent to a solution containing 5.0 gram NaCl per litre, in so far as electrolyte osmotic action is concerned, shall not produce a haemolysis value higher than 10% and a salt solution of 4.0 gram per litre shall not differ by more than 10% in haemolysis value from that caused by the corresponding control solution.

(b) Method:

From the primary buffer stock solution for haemolysis three solutions are prepared: 30 ml buffer stock solution and 10 ml water (solution a0), 30 ml buffer stock solution and 20 ml water (solution b0) and 15 ml buffer stock solution and 85 ml water (solution C0).

To each of three centrifuge tubes (1, 2 and 3), 1.40 ml extract are added. To tube 1 is added 0.10 ml a0, to tube 2, 0.10 ml b0 and to tube 3, 0.10 ml C0, thus obtaining salt solutions equivalent to solutions containing 5.0 (tube 1), 4.0 (tube 2) and 1.0 gram NaCl per litre (tube 3) in so far as electrolyte osmotic action is concerned. To each tube is added 20 µl fresh, well mixed heparinised human blood. The tubes are put into a water bath at 30° C (± 1° C) for 40 minutes. Then three solutions containing 3.0 ml a0 and 12.0 ml water (solution a1) 4.0 ml b0 and 11.0 ml water (solution b1), and 4.75 ml b0 and 10.25 ml water (solution c1) are prepared.

To the first tube is added 1.50 ml of a1, to the second 1.50 ml of b1 and to the third 1.50 ml of c1. The tubes are centrifuged for 5 minutes at 2,000 to 2,500 rpm in a swing-out centrifuge. Concurrently, control solutions, in which the extract is replaced with water, are prepared for each of the concentrations.

The extinction at 540 nm of the liquid layer is measured. Buffer stock solution for haemolysis is used as blank. The haemolysis value in per cent is calculated according to the following formula:

where E100%= extinction for the solution containing an equivalent of 1.0 gram salt per litre

and Eexp = extinction for the solutions containing an equivalent of 4.0 and 5.0 gram salt per litre respectively

Buffer stock solution for haemolysis

90.0 g sodium chloride, 13,7 g anhydrous disodium phosphate and 1.90 g anhydrous monosodium phosphate are dissolved in distilled water and made up to 1000.0 ml.

Regeling
Protocol bij de Europese Overeenkomst betreffende de uitwisseling van geneesmiddelen van menselijke oorsprong, zoals gewijzigd
Soort
Verdrag
Geldend vanaf
19-04-1982
BWB-id
BWBV0002901
Versie
1982-04-19_0

In de hele regeling · Officiële tekst op wetten.overheid.nl