- Prepared, sterile and tested solution of sodium diatrizoate endotoxins and polysaccharide
- Suitable for the isolation of human polymorphonuclear leukocytes (granulocytes) from undiluted whole blood
Intended Use: For Research Only
Polymorphprep™ is a ready-to-use, sterile, endotoxin-tested solution for the isolation of polymorphonuclear leukocytes (PMNs) from undiluted human whole blood. The solution contains sodium diatrizoate and a polysaccharide
Density: 1.113 +/- 0.001 g/ml
Composition: Sodium diatrizoate 13.8% (w/v), polysaccharide 8.0% (w/v)
Osmolality: 440-500 mOsmol/kg
Endotoxin: < 1.0 EU/ml
Stability and Storage
Polymorphprep™ is stable for 3 years provided the solution is kept sterile and protected from direct sunlight. Prolonged exposure to direct sunlight causes the release of iodine from the sodium diatrizoate molecule. This effect is negligible when working with this solution on a day-to-day basis. Polymorphprep™ should be stored at room temperature (+4°C to +30°C).
With the exception of basophils, polymorphonuclear leukocytes (PMNs) have a much higher buoyant density than mononuclear cells, >1.085 g/mL. Unfortunately, the buoyant density of erythrocytes tends to be 1.09-1.11 g/mL, making it difficult to separate from whole blood using a density barrier similar to that used for mononuclear cells. A number of procedures have been developed in an effort to overcome these difficulties.
The high osmolality of Polymorphprep causes red blood cells to lose water and shrink, thereby increasing their effective buoyant density. This allows the dextran-aggregated erythrocytes to rapidly sediment through the dense medium. Because the osmotic gradient between the medium and the erythrocytes decreases as the cells sediment more into the medium (i.e., the water loss from the erythrocytes is greatest at the top of the PolymorphprepTM and decreases progressively as they sediment more). ), a diatrizoate gradient forms within the density barrier.
PMNs form a band within this density gradient while mononuclear cells remain at the sample/medium interface. The method is effective only with undiluted whole blood, not with a leukocyte-rich fraction. Temperature is important for optimal results, as changes in temperature affect the density and viscosity of the PolymorphprepTM solution. The temperature of the blood sample and the medium must be maintained between 18 and 22 degrees Celsius.
Analysis of the upper and lower bands from the PolymorphprepTM separation using a Coulter STKR cell analyzer is shown in the figure to the right. The analyzer determines the number of cells in the sample (ordinate) based on cell volume (abscissa). The relative number of cells is the number of cells in a particular volume expressed as a fraction of the total in each sample. The upper band contains only lymphocytes (40-80 fl) and monocytes (80-130 fl); all PMNs (150-320 fl) are in the lower band which has negligible mononuclear cell contamination. Contamination of the PMN band by erythrocytes is between 2-6% of the total number of cells.