Lab 2 ; Nur'Ain binti Zulkefli- 111399

LAB 2: MEASUREMENT AND COUNTING OF CELLS USING MICROSCOPE

2.1 Ocular Micrometer

   Introduction

     Ocular micrometer is use in order to measure and compare the size of prokaryotic and eukaryotic microorganism. Microorganisms are measured with an ocular micrometer which is inserted into the one of the microscope eyepieces. The micrometer, which serves as a 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 the eyepieces, 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 10 µm apart etched. By determining how many units of the ocular micrometer superimpose a known distance on the stage micrometer, you can calculate the exact distance each ocular each ocular division measures on the microscopic field. When you change objectives you must recalibrate the system. After calibration of ocular micrometer, the stage micrometer is replaced with a slide containing microorganism. The dimensions of the cells may then be determined.

   Objective

To measure and count cells using microscope

     Results

1)     Lactobacillus

    40 x magnifications

100 x magnifications

2)   Yeast

40 x magnifications

100 x magnifications

          Discussion

To compare and measure the size of prokaryotic and eukaryotic microorganisms, we used ocular micrometer. We use the lowest power objective to focus the microscope until the image on the stage micrometer is superimposed on the eyepiece scale and calibrate.

Prokaryotic and eukaryotic has different characteristic. Prokaryotic cell is a single cell while eukaryotic cell is more complex than prokaryotic and can be single or multicellular  cell. Prokaryotic cell has smaller size because prokaryotic does not contain many organelles compared to eukaryotic such as chloroplast, nucleus, endoplasmic reticulum and cell membrane.

The example of prokaryotic cell in this experiment is lactobacillus and yeast as the example of eukaryotic. During the experiment, we used  40x magnification and 100x magnification to observe the both cells.

	400x magnification		1000x magnification
calibrate	Stage micrometer	Ocular division	Stage micrometer	Ocular divison
	0.03 mm	38 division	0.09 mm	96 division
	7.9x10⁻⁴ mm	1 division	9.38x10⁻⁴ mm	1 division
	0.79 µm	1 division	0.94 µm	1 division

Size of microorganism :                                            2 division                                               5 division

YEAST :

400x magnification

       4 division x 0.79 µm

       =3.16 µm

1000x magnification

       7 division x 0.94 µm

       = 6.58 µm

LACTOBACILLUS :

400x magnification

       2 division x 0.79 µm

       =1.58 µm

1000x magnification

      5 division x 0.94 µm

      =4.7 µm




    Conclusion

As the conclusion, we are able to learn on measuring and counting the cells using microscope. An ocular micrometer is a glass disk that attaches to a microscope’s eyepiece. An ocular micrometer has a ruler; calibrated with a slide micrometer that allows the user to measure the size of magnified objects. The distance between the marks on the ruler depends upon the degree of magnification. We used this ocular micrometer to observe prokaryotic and eukaryotic cells since these two cells are very small.

Reference

http://wiki.answers.com/Q/What_is_ocular_micrometer

http://www.ehow.com/how_5019336_use-ocular-micrometer.html

Pearson International edition Biology 8th Edition, Campbell. Reece

2.2 Neubauer Chamber

Introduction

          Neubauer chambers are more convenient for counting microbes. The Neubauer is a heavy glass slide with two counting areas separated by a H-shaped trough (see figure 2.1). A special coverslip is placed over the counting areas and sits a precise distance above them.

Objective

To count cells using microscopes.

Result

Yeast 400x magnification

Discussion

          


          In the second experiment, Neubauer chambers are used for more accurate in counting microbes. In addition, the sterilization method is used to avoid contamination of microorganism. The types of sterilization that we used is flame sterilization and also Pasteur pipette to transfer the yeast to the slide.

          The chamber consists of 9 major large squares. In the calculation, we only use the middle of the large square.  Inside the large square, there contain another 16 smaller squares. Then choose 10 squares out of 16 squares to calculate the number of yeast.

56 ÷ 10 = 5.6

                 = 6

Volume = 0.05 x 0.05 x 0.01 mm

               = 2.5 x 10⁻⁵ mm

               = 2.5 x 10⁻⁷ ml

6 cells in 16 squares

1 square = 6 ÷16

                 = 0.375 cells in 2.5 x 10⁻⁷ ml  

1ml = 0.375 ÷ 2.5 x 10⁻⁷

       = 1500000 cells/ml

Conclusion

          As the conclusion, we can use two methods to measure and count cells using methods. The first one is by using ocular micrometer and the second one by using Neubauer chambers. By using Neubauer chambers, we can get more convenient in counting the cells compared to ocular micrometer. This is because, when using Neubauer chambers, the sterilization is needed to avoid any contaminant. Hence,  Neubauer chambers are suitable to count the cells.

Reference

  1.      Pearson International Edition Biology, 8^th Edition, Campbell, Reece