Plasmodium Falciparum is a human parasite that lives exclusively within red blood cells. In the process of growing, it uses the molecules within the erythrocytes to feed off them, and since the molecules of iron within the hemoglobin would be toxic to the parasite, it takes them and places them into an inert crystal called hemin. This crystal of hemo confers some param magnetism to the parasites.
It is this paramagnetic property that is used to separate infected erythrocytes from those that do not carry a parasite inside. With the use of a magnet, a culture that has very low percentage of parasites can be concentrated to achieve almost a hundred percent paras. This separation of infected from uninfected red blood cells is very desirable in a number of experiments where a particular trait of the parasite needs to be studied without the confounding factors of surplus erythrocytes.
The methodology shown here is fast, uncomplicated, and harmless to the parasite. Today We will be describing the procedure of plasmodium falciparum separation of chizen stage. This multi-component equipment has to be assembled for every chant.Isolation.
First place the column holder on the metallic stand, and then the magnetic column is placed on set holder. For our procedure, we use Ls, columns of milian. Next, we use a 50 ml tube to add 23.2 mls of RPMI 1640, and 750 microliters of a 7.5%sodium bar carbonate solution, which is to be the work solution for this process to be successful, it is necessary to calibrate the column Before we start.
For this, add three mls of the work solution and this will flow into the discard labeled 50 ml Tube. Then add eight mls of the plasmodium FCI burn culture to the column. Watching that as soon as the last part of the work solution has been pushed out of the column by the culture, the tube be changed to the other 50 ml tube below.
To begin collecting the Culture. Once the flow of The culture through the column has ceased, add one ml of the work solution to the column to push the remaining unattached red blood cells. Out of the next, add three mls of the work solution to the column and switch to the discard tube so as not to alter the proportions of the culture.
Once The three mls have gone through the column, detach it from the stand and place any 15 ml tube. Next, add three mls of the work solution. This will elute the chizen that have been trapped in the beads of the column.
Depending on the experiment that is being performed, we are able to collect as many parasites as we need. If our culture has a high semia, we may obtain the amount of chant needed in the first collection. If not, since the binding capacity of the column is probably less than our shant content, we can pass our culture through the column more times and collect more parasites.
After this is done, we centrifuge the 15 ml tube for five minutes at 150 centrifugal forces. After centrifugation, we may see the pellet formed at the bottom of the tube. We then remove the SUP natant working solution, leaving enough to make it possible to determine our yield through microscopy.
We can then decide whether to repeat or stop the collection at that point, and if so, collect more chizen in the same 15 ml tube. This Procedure can be applied for invasion essays, synchronization of the parasites, and growth essays.
