LAB 1 : PRINCILES AND USE OF MICROSCOPE

Laboratory's Information :
Laboratory's Number - LAB A203
Laboratory Title : PRINCIPLES AND USE OF MICROSCOPE
Lecturer's Name : Associate Professor Dr. Liong Min Tze
Laboratory Assistant : Madam Najmah
Group Members : Charles Ng Wai Chun, Siti Hawa Binti Ramli, Nuramirah Binti Ramlan
Date of Laboratory : 22 September 2015

LAB 1 : PRINCILES AND USE OF MICROSCOPE

1.1 SETTING UP AND USING THE MICROSCOPE

Introduction

            A microscope is an instrument used to see objects that are too small and to enlarge images of small objects that cannot be seen by naked eyes. The science of investigating small objects using such as instrument is called microscopy. “Micro” refers to tiny while “scope” refers to view. There are many types of microscope such as electron microscope, the ultramicroscope and the various types of scanning probe microscope. The most common is the optical microscope, which uses light to image the sample.

The Optical Microscope




Parts and Function of an Optiocal Microscope

Eyepiece	Contains the ocular lens, which provides a magnification power of 10x to 15x, usually. This is where you look through.
Nosepiece	Holds the objective lenses and can be rotated easily to change magnification.
Objective lenses	Usually, there are three or four objective lenses on a microscope, consisting of 4x, 10x, 40x and 100x magnification powers. In order to obtain the total magnification of an image, you need to multiply the eyepiece lens power by the objective lens power. So, if you couple a 10x eyepiece lens with a 40x objective lens, the total magnification is of 10 x 40 = 400 times.
Stage clips	Hold the slide in place.
Stage	It is a flat platform that supports the slide being analyzed.
Diaphragm	It controls the intensity and size of the cone light projected on the specimen. As a rule of thumb, the more transparent the specimen, less light is required.
Light Source	It projects light upwards through the diaphragm, slide and lenses.
Base	Supports the microscope.
Condenser lens	It helps to focus the light onto the sample analyzed. They are particularly helpful when coupled with the highest objective lens.
Arm	Supports the microscope when carried.
Coarse Adjustment Knob	When the knob is turned, the stage moves up or down, in order to coarse adjust the focus.
Fine Adjustment Knob	Used fine adjust the focus.

Magnification and Resolution

1) Basically, the microscope that we used in the laboratory has four different magnifications of objective lenses, which are, 4x objective lens, 10x objective lens, 40x objective lens and 100x objective lens which is also called oil-immersion lens.
2) The total magnification of the images that we will be seeing from the microscope is calculated by multiplying the objective lens power by the eyepiece lens power.
- The total magnification is calculated as follows :
i) 4x objective lens power X 10x eyepiece lens power = 40x magnification
ii) 10x objective lens power X 10x eyepiece lens power = 100x magnification
iii) 40x objective lens power X 10x eyepiece lens power = 400x magnification
iv) 100x objective lens power X 10x eyepiece lens power = 1000x magnification    
   
3) The ability to distinguish two very small and closely spaced object as separate entities known as resolution or resolving power. 
4) Closing of the diaphragm will cause the image contrast but decrease in its resolution while the opening of the diaphragm will decrease the contrast but increase its resolution. 
5) Ergo, the magnification and resolution are greatly related to each other and both of them are important in achieving a clear image that we wanted to see from a slide. 

         

                                           4x, 10x, 40x and 100x Objective lenses.

           10x Eyepiece lens. 

Objective

1) To know the ways to use and take care of a microscope correctly and properly.

2) To learn the importance of magnification and resolution of the microscope

Materials and Reagents

1) Microscope slide.

2) Cover slip.

Procedure

(refer to the laboratory manual)

Results

Lactobacillus under the 4x objective lens (40x magnification). 

    

 Lactobacillus under the 10x objective lens (100x magnification). 

                 

                                           

Lactobacillus under the 40x objective lens (400x magnification). 

    

  Lactobacillus under the 100x objective lens (1000x magnification). 

Discussion

1) When the Lactobacillus slide was observed under the 4x objective lens which is 40x magnification, we can only see the bacteria in a very small form which cannot be seen clearly.
2) When the Lactobacillus slide was observed under the 10x objective lens which is 100x magnification, the image formed is bigger than the previous objective lens and it is clearer as well.
3) When the Lactobacillus slide was observed under the 40x objective lens which is 400x magnification, we can roughly see the outline of the bacteria in a much more clearer form of image.
4) When the Lactobacillus slide was observed under the 100x objective lens (oil-immersion lens) which is 1000x magnification, the shape of the Lactobacillus can be seen and examined clearly. We can still roughly distinguish the bacteria even though it was overlapped. However, an electron microscope will be needed if we want to distinguish every bacteria in a slide.
5) Scientific classification of Lactobacillus :

    Scientific name : Lactobacillus
    Rank : Genus  
    Kingdom : Monera
    Division : Firmicutes
    Class : Bacilli
    Order : Lactobacillales
    Family : Lactobacillaceae

    Morphology of lactobacillus :
    - rod-shaped
    - convex
    - smooth
    - opaque without pigments

Conclusion

            Microorganisms are small living organism that needed to be magnified in order to be seen. Magnification helps us to see the objects seems bigger and meanwhile, resolution helps us to distinguish the two things from each others.

            Microscope is a very important instrument in aiding us seeing and examining the cells which cannot be seen with our naked eyes. And here are some precautions and steps that should be taken during the experiment:

1) Microscope is an expensive instrument which we need to handle it very carefully.               

2) When we are to examine a glass slide, always begin with the scanning (4x objective) or the low power objective (10x). Never begin with the high power objective (40X). It will be difficult to find what we are looking for under high power.

3) For better result, turning the coarse adjustment until the stage is as close to the objective as it will go.

4) Always clean the slides and microscope when finished the experiment. Be careful when handling glass slides and cover-slips.

Reference

1) Classification of Lactobacillus Acidophilus. Retrieved from 

http://cahoney-l-acidophilus.pbworks.com/w/page/6327452/Classification

2) Matthew K.Nelson (2011), Safe / Use of the microscope / Cladistics. Retrieved from http://www.natureboy.com/3454/labexercise1.pdf 

3) R. Bunemann & G. Wiliams (2008), Lactobacillus characteristics. Retrieved from http://www.coloss.org/beebook/I/gut-symbionts/4/4/2

4) Wikipedia, the free encyclopedia (3 October 2015), Microscope. Retrieved 

from https://en.wikipedia.org/wiki/Microscope

5) Wikipedia, the free encyclopedia (2 October 2015), Lactobacillus. Retrieved from https://en.wikipedia.org/wiki/Lactobacillus

1.2 EXAMINATION OF CELLS

Introduction

            Some if the specimens of bacteria cannot be identified directly under the microscope because of their extremely small in size. The use of low-power of high-power objective lenses are far not enough for the studying of the bacteria. In addition, staining or drying the cells for the studying of microorganisms will definitely distort them in terms of shapes and sizes, which eventually destroy them. Therefore, some specimens need to be suspended in a drop of liquid (such as immersion oil) for microscopic examination to fill the space between the cover slip and the slide which it allows light pass through it easily. Nonetheless, we can even determine whether the bacteria or cells are motile with this technique. And, this is known as “wet mount” technique. 

Objective

1) To provide an experience in the use of microscope.

2) To illustrate the diversity of cells and microorganisms.

Materials and Reagents

1) Culture

2) immersion oil

3) Lens tissue

4) A microscope slide containing stained microorganisms

5) Inoculating loop

6) Bunsen burner

7) Slide and coverslip

Procedure

(refer to the laboratory manual)

Results

Lactobacillus wet mount under the 100x objective lens (1000x magnification).

 Discussion

1) Under the 100x objective lens of 1000x magnification, we could see the shape of Lactobacillus which is rod-shaped. 

2) It can be clearly seen that the Lactobacillus was moving around the glass slide as well which proved its motility and was still alive.  

3) Lactobacillus is any of a group of rod-shaped, gram-positive, non-spore-forming bacteria of the family Lactobacillaceae.

4) Lactobacillus is a facultative anaerobe which they can produce energy through glycolysis and fermentation when oxygen is not present. 

5) Similar to other genera in the family, Lactobacillus is characterized by their ability to produce lactic acid as a by-product of glucose metabolism in its metabolism system.

6) Lactobacillus is generally non - motile and can survive in both aerobic and anaerobic environments. 

7) Lactobacillus is also a type of "friendly" bacteria that normally live in our digestive, urinary, and genital systems without causing disease. Lactobacillus is also used in some fermented foods like yogurt and in dietary supplements.

8) Commercial preparations of lactobacilli are used as probiotics to restore normal flora after the imbalance created by antibioric therapy.

Conclusion

            Based on the experiment we conducted, we found that using the wet mount technique with the aid of 100x objective lens or oil-immersion lens, we could see and examine the bacteria clearly including its size and shape. And from the experiment, we got to know that the Lactobacillus is a motile microorganism organism that have a rod-shape. This could be done because no staining or drying of specimens is needed and hence it maintain the shape and size of the bacteria without destroy it. Last but not least, to achieve the best condition of the specimens for the purpose of study, we must practise the aseptic technique during the preparation of the specimens in order to prevent contamination of the slides which may cause different results in our experiments. 

Reference

1) Oliver Kim (2008 - 2014), Making a wet mount microscope slide. Retrieved from http://www.microbehunter.com/making-a-wet-mount-microscope-slide/

2) David B. Frankhauser, Ph.D. (24 June 2005), PREPARATION OF WET MOUNT SLIDE. Retrieved from http://biology.clc.uc.edu/fankhauser/Labs/Microbiology/Wet_Mount/Wet_Mount.htm 

3) Wikipedia, the free encyclopedia (2 October 2015), Lactobacillus. Retrieved from https://en.wikipedia.org/wiki/Lactobacillus

4) WebMD (2005 - 2015), Find a vitamin or supplement LACTOBACILLUS. Retrieved from https://en.wikipedia.org/wiki/Lactobacillus

5) The Editors of Encyclopedia Britannica (2005), Lactobacillus bacteria. Retrieved from http://global.britannica.com/science/Lactobacillus