I am currently doing a study in water quality of a lake and need to calculate the total number of cells of phytoplankton in a sample, I already have the no of cells counted per view, no.of field of views. Could anybody help out please?
You can use hemocytometer, but the problem is, if the plankton cells are big then it is hard to accommodate them in the chamber. If you use the old model neubauer hemocytometer, where you need to place a cover slip on top of the sample which will provide more room and even if the cells are big, you should be able to count them.
Cells/mL = Total number of cells counted/number of field of view X DF X CF
Let the phytoplankton to settle in a settling chamber. Then, using an inverted microscope (conventionally) they could be enumerated. You need to pre-determine the sample volume required for the analysis. Most samples are settled at 25 mL. When samples are difficult to count are 5 mL or 2.5 mL sample volumes used. The 50 mL samples are used when very low number of phytoplankton are present in the samples. please note that only chloroplast containing phytoplankton are counted. Report the results of the sample sedimentation procedure as cells/mL, which is calculated as:
cells/mL = cell count X total area of chamber bottom / Length of strip of the counting chamber X Width of strip of the counting chamber X Volume of the counting chamber X No. of strips counted.
Alternatively, you can use a haemocytometer (as suggested above) or a flow-cytometer or a fluorimeter for the purpose.
You can count the cell in 1ml sample by using SR chamber under 200X. And then calculate to XX cells/ml or XX cells/L. Good luck.
A compromise counting cell is the Palmer plankton counting chamber. Its depth and volume are nearly optimal for phytoplankton counts. I fill the chamber with about 200 microliters of the sample and count at 500X.
If the sample is of insufficient population density, then an inverted microscope and a settling column is helpful. If you don't have an inverted 'scope, then you can take a large volume of sample water and allow the cells to settle to concentrate them or you can concentrate them with centrifugation.
If you use a centrifuge, there is a continuous centrifugation apparatus invented by Albert Szent-Györgyi but those are rare these days and somewhat temperamental to use.
1) it is necessary to know the volume (ml) of sample
2) stir the water in the sample
3) Remove 1 ml of water with phytoplankton and placed on a glass slide
4) under a microscope to count the number of cells
5) do a selection of 5-10 times,
6) to determine the average with an error
7) counted on the sample volume
You have to concentrate the sample via sedimentation or filtration. might be the following link is useful:
http://www.epa.gov/grtlakes/lmmb/methods/phy.pdf
The objective needs to be clarified. Phytoplankton size range starts from picoplankton,
Re: Pengbin Wang's answer - "SR" = "Sedgewick Rafter" chamber
The method used will depend on the expected cell density and size classes. Sedgewick Rafter chambers should be satisfactory and will give you a greater volumn than Neubauer hemocytometer. Samples will need to be preserved with Lugol's Iodine solution or the like. If not much material is visible you can preconcentrate your samples or, if you have access to an inverted microscope, use a Utermoehl chamber.
There are plenty of instructions on the web on how to use the chambers mentioned above.
I suppose you did the counting using an inverted microscope and a sedimentation chamber. If you already have the no of cells counted per view and the no.of field of views, to calculate the number of cells in your sample you need other two parameters: the total volume of the sedimentation chamber (= volume of sample counted) and the total number of the microscopic fields corresponding to the bottom of the sedimentation chamber. You can estimate this last number by measuring the area of a microscopic field, using a micrometer eyepiece (be aware that this value is dependent on the objective you use) and the bottom area of the sedimentation chamber. The ratio between the bottom area of the chamber and the area of a microscopic fields is equal to the total number of counting fields in your chamber. Now you can calculate the cell number as follows:
no. of cells/ml = (total number of cells counted * total number of microscopic fields in the chamber/number of fields analysed) / volume in ml of the chamber
Thank you all for all the answers. They were all quite helpful. I have been able to calculate the total phytoplankton in the water sample.
sedgewich rafter Chamber is the best method to count the Phytoplankton.
Seasonal Abundance of Micro Algae in Pandi Backwaters of Godavari Estuary, Andhra Pradesh, India
Notulae Scientia Biologicae. 2010;2(3)26-29 DOI 10.15835/nsb.2.3.4685
check this publication for counting details.
Cell counting with Sedgwick rafter counter is the standard method for counting of phytoplanktons as well as zooplanktons. It's a very easy and handy method for both the quantitative and qualitative assessment. You may visit the following web link:
http://courses.washington.edu/uwtoce12/methods/sops/phytoplankton.pdf
Dear friend,
How to calculate concentration factor for net collected phytoplanton sample
http://nio.org/userfiles/file/biology/Phytoplankton-manual.pdf found this to be useful.
Use Sedgewick rafter cell method, follow the guideline commented above. you will be benefited.
To calculate the total phytoplankton cells use standard method by using Sedgewick Rafter (S-R) cell.
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