The present investigation concerns a device representing a new kind of liquid flow system in connection with liquid flow cytophotometers.
Cytophotometry is an important operating method in cell biology research, especially in the field of comprehensive cancer research. Thus, it is considered important to investigate the abnormal growth in cell populations where new cells are created by means of cell division. During its synthesis phase the cell will double its DNA-content. In this way one gets two new cells when the cell divides (the mitotic phase). By measuring the DNA-content per cell it is possible to determine the distribution of the cells on the various cell cycle phases. The fraction of cells containing an increased DNA-content in a cell population can be a measure of the multiplying activity. It is important that the distribution of the cells in a cell population on the various cell cycle phases can be determined with a great accuracy. Statistical errors can only be avoided by measuring several thousand cells. This necessitates that the individual measurements are done very rapidly.
The so-called liquid flow cytophotometry is a technique which is now applied for such measurements. Today there is an increasing use of this technique in connection with research as well as diagnostic purposes. The principle of such a method is to lead a limited flow of cells stained with a fluorescent dye, quantitatively bound to the cell components which are to be studied, through a beam of exciting light and then measure the intensity of the resulting fluorescence pulses. The distribution of the cells with regard to a certain component, such as DNA, can thus be determined with great accuracy and with a rate of the order of 10.sup.3 per second. The cells are automatically transported to the measuring spot in a liquid flow, usually water.
Thus, the cells are not spread on a glass plate, but are kept in a liquid suspension. This suspension is driven through a capillary tube, and by means of a concentric sheath flow flowing around the tube mentioned above and in the same direction as the cells and containing no particles, the cell flow is hydrodynamically focussed so that the cells pass by through a strictly limited area in the cytophotometer's measuring focus. The cells pass by one after another through this measuring area at a high speed, just like mentioned above.
The measuring of the cell components is done by means of a microfluorometer. As mentioned above, the cell component must be stained with a fluorescent dye. When a cell containing such a fluorescent dye passes through the microscope lens' focussing area, it excites fluorescent light which is accumulated by the lens and then lead to a photomultiplier. The signal from this photomultiplier is registered and expresses the cell's contents of the component in question.
A well-known liquid flow cytophotometer applies the technique mentioned above. It comprises a closed system including a sheath flow measuring chamber. This chamber consists of a glass- or metal body with channels in a T-form. By means of suction the sheath flow and the particle suspension which is to be measured are led through the channel forming the T-form's system, and accurately centered along the system's optical axis. In this way one gets a measuring area at the T-form's peak, i.e. at the area of the perpendicular channels constituting the T-form's arms. One of these perpendicular channels transports a cleansing agent, while the other represents a drainage channel for the fluid flow containing measured particles. Thus, by using this well-known technique one gets a supply of particles along the optical axis with a focussing of every particle through the optical focus lying on the measuring aperture level. The optical system which is applied here comprises a so-called oil immersion optics which is known to the expert. The optics is in contact with the liquid suspension containing the particles or the cells which are to be studied.
There are liquid flow cytophotometers using closed sheath flow measuring chambers, but where the liquid flow is led perpendicularly through the exciting light in the measuring area.
Further, there are liquid flow cytophotometers where a focussed liquid jet containing particles which are to be studied, is led--in the air--through an excitation light falling on the liquid jet. This light is focussed in the liquid jet, so that when single particles, which can be stained with a fluorescent dye, pass through this light focus, one will have a pulse of fluorescent light which is picked up by a photo cell.
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