Interesting, can you provide more information such as brand, model and how it´s work? It´s for count parasites in culture? or in after a blood smear? I never work with such equipment but I have interest in know more about it. Thanks

I use Coulter Counter Z1 series (www.beckmancoulter.com/). It works well for the extracellular parasite, Trypanosoma brucei. It will probably not work if there are other types of cells in culture (blood for example).

Contact the companies, have them come into your lab and demonstrate their machines on your samples is the best way to select equipment to purchase

Hi Yael and all

In my PhD project, I had used the CASY cell counter and analyser system (Innovatis AG, Germany) to count Trypanosoma brucei cells. It had worked great which give me cell number, sizes and even nuclear DNA content profiles and dead cell data. Though we never tried on on any other cell line like Leishmania but very easy to contact equipment distributer and ask.

Hope this will help

Best

My lab uses a Casy Cell Counter (Scharfe) http://www.rjmsales.com/casy.htm

We count bloodstream and procyclic Trypanosoma brucei; promastigote and amastigote forms of Leishmania parasites; and epimastigote and trypomastigote forms of Trypanosoma cruzi. The instrument is not very expensive, robust and reliable with a small footprint if you want to keep it in a biosafety lab.

If you are going to count epimastigote forms in T. cruzi or promastigote in Leishmania species a electronic counter works. However I do not think the same for counting amastigote

Since both parasites types (Leishmania and Trypanosoma) have a wide morphological spectrum into each step of the life cycle (tripo, epi or amatisgotes) I will recommend you to explore the flow cytometry as a alternative way. It more expensive but you will get information of high quality. Also, you probably can identify the parasites in presence of other cellular types. Obviuosly, I dont know your experimental model, so these observations could be not useful.

Possible by microscope see all stage of flagellate until cyst shape to parasite

Intracellular RBC parasites, and perhaps extracellular parasites may be labeled with fluorescent dye (acridine orange) and sorted based upon cellular fluorescence. This is useful in mammalian blood, as RBC do not normally contain nucleic acids. Flow cytometer could separate fluorescent cells based upon size and fluorescence intensity.

Hello Yael,

I worked for many years with T.cruzi and more recently with Leishmania. If you are thinking of using the counter to count T. cruzi in blood, I would advise you to try to get more information from Alain Fairlamb and Mohamed Bessat who also answered your question.

In my experience automated counters can be useful when you have a suspension of parasites whether from liquid culture ( T. cruzi epimastigotes or trypomastigotes or even transformed amastigotes or Leishmania promastigotes-possibly also metacyclics) . It could also work for intracellular amastigotes after lysis of host cells, if (and that is a problem) you have removed the majority of other contaminating cells and debris from the suspension.

For counting T.cruzi trypomastigotes in blood where the frequency of parasites to blood cells is very low even in heavy parasitemias, the counting machines could not distinguish the parasites from cells of similar size and shape or debris However, better machines may have appeared in the last 10 years).

There are ways to get rid of RBCs and platelets. For instance, red cells can be lysed with hypotonic solution and Ca: the debris, aggregated platelets and WBC are allowed to sediment at 1 g for 20 min at RT. The living trypos will swim to the surface and can be sampled. This and other methods have been published around 1970-80, but in the end they turned out to be more cumbersome and time consuming than traditionally counting of parasites in a fresh blood smear.

Ises A. Abrahamsohn

University of São Paulo, Brazil.

I agree with Abrahamsohn. For T. evansi and T. vivax trypomastigotes counting in fresh blood smears seens more acurate than automatic methods. An alternative to the use of lysing solution is centrifugation of whole blood in Percol (1:1).

Fabiano Cadioli, Kindly shed more light on your reply, I'm interested in the formulation of lysing solution and Percol

Rolayo Ogunwole, the use of Percol was decribed in GONZÁLES, L. F.; GARCÍA, J. A.; NÚÑEZ, C.; PERRONE, T. M.; GONZÁLEZ-BARADAT, B.; GONZATTI, M. I.; REYNA-BELLO, A. Trypanossoma vivax: A novel method for purification from experimentally infected sheep blood. Experimental Parasitology. v. 111, n. 2, p. 126-129, 2005.

I add lysis solution for commercial blood sample, wait a minute (2 minutes) and make a blood smear staining with Rosenfeld. This technique is best for samples with low parasitemia and fibrin clots (or roleaux).

Here is a link for RBC lysis solution:

http://depts.washington.edu/flowlab/Cell%20Analysis%20Facility/RBC%20Lysing%20Solutions%20and%20Cell%20Lysing%20Procedure.pdf

Good work!

Fabiano Cadioli, many thanks.