No. Matrice : 111410
2.1 Ocular Micrometer
Introduction :
Ocular micrometer is use in order to measure and compare the size of prokaryotic and eukaryotic microorganisms. Microorganisms are measured with an ocular micrometer which is inserted into the one of the microscope eyepieces. The micrometer, which serves as scale or rule, is a flat circle of glass upon which are etched equally spaced divisions. This is not calibrated, and may be used at several magnifications. When placed in a eyepiece, the line superimposed certain distance markers on the microscope field. The actual distance superimposed may be calibrated using a stage micrometer on which parallel lines exactly 10micrometer apart etched. By determining how many units of the ocular micrometer superimpoe a known distance on the stage micrometer, you can calculate the exact distance each ocular division measures on the microscopic field. When you change objectives you must recalibrate the system. After calibration of the ocular micrometer, the stage micrometer is replaced with a slide containing microorganisms. The dimensions of the cells may then be determined.
Objective :
To measure cells using a microscope.
Result :
1. With 40x objective len, each division of ocular micrometer :
(1x0.01) / 4 = 0.0025mm
(a) Saccharomyces cerevisiae (Yeast)
2 x (0.0025) = 0.005mm
(b) Lactobacillus fermentum
1 x 0.0025 = 0.0025mm
2.With 100x objective len, each division of ocular micrometer :
(1x0.01) / 10 = 0.001mm
(a) Saccharomyces cerevisiae (Yeast)
5 x (0.001) = 0.005mm
(b)Lactobacillus fermentum
2.5 x 0.001 = 0.0025mm
Discussion :
1.An ocular micrometer has a ruler that allows the user to measure the size of magnified objects.
2.Ocular micrometer can be used to measure any planar dimension in a microscope field since the
ocular can be turned in any direction and the object of interest can be repositioned with the stage
manipulators.
3.With difference magnification, the division of ocular micrometer will has difference value. Thus, it
is necessary to calibrate the scale by focusing on a fix and known scale micrometer (a stage
micrometer) placed directly on the stage for every magnification.
4.After calibrated, the ocular micrometer can be use to measure the dimension of cells.
Conclusion :
Ocular micrometer with a known scale stage micrometer , the cell can be measure easily and accurately especially with oil immersion method.
Reference :
1.How to Use an Ocular Micrometer | eHow.com
2.http://www.ehow.com/how_5019336_use-ocular-micrometer.html#ixzz1rQeXORkR
3.http://www.ruf.rice.edu/~bioslabs/methods/microscopy/measuring.html
2.2 Neubauer Chamber
Introduction :
Neubauer chambers are more convenient for counting microbes. The Neubauer is heavy glass slide with two counting areas separated by a H-shaped trough. A special coverslip is placed over the counting areas and sits precise distance above them.
Objective :
To count cells using a microscope
Result :
Observation of yeast on Neubauer chamber under 40x objective magnification.
1. Volume of one small box :
Height = 0.2mm
Length = 0.2mm
Breadth = 0.1mm
- 0.2mm x 0.2 mm x 0.1mm = 0.004mm^3
= 4x10^-6 cm^3
= 4x10^-6 mL
2. Total no. of cells calculated from 10 small box :
9+14+8+13+22+11+20+15+5+7 = 124
- average = 124 / 10
= 12.4
3. Concentration of cell (no. of cells / each mL) :
- 12.4 / (4.0x10^-6)mL = 3100000 cells / mL
Discussion :
1.The Neubauer chamber is a device used for determining the number of cells per unit volume of a
suspension.
2.Coverslips for counting chambers are specially made and are thicker than those for conventional
microscopy, since they must be heavy enough to overcome the surface tension of a drop of liquid.
3.Suspensions should be dilute enough so that the cells or other particles do not overlap each other on
the grid, and should be uniformly distributed.
4.When counting the cells, cells that overlap a ruling, count a cell as "in" if it overlaps the top or right
ruling, and "out" if it overlaps the bottom or left ruling.
5.Counting :
(a) The chamber contains many grids, producing nine major large squares.
(b) For calculation purposes, only the middle large square which has 16 small squares with area
(0.2mm x 0.2mm) is used.
(c) Choose randomly 10 small boxes and count the total number of cells and its average per square.
(d) Assuming the average number of cells = Z, and volume of a square = 4x10^-6mL.
Then the concentration of cells = Z / (4x10^-6) cells/mL.
Conclusion :
Neubauer Chamber is a useful advice use to count concentration of the cell by calculate the average of cells in a sample on the small boxes of chamber which chosed randomly and divided to the volume of each box.
Reference :
1. http://www.ruf.rice.edu/~bioslabs/methods/microscopy/cellcounting.html
2.http://www.uni-greifswald.de/~immuteach/methods/counting_chamber/counting_chamber.html#
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