Micrometry and Microscopy

Abstract

There are organisms that are too small to measure so there are certain equipment used  to determine their exact size. This exercise aimed to measure the specimen Didinium nasutum using the concept of micrometry and microscopy with the aid of  a compound microscope. The calibration constant of low power objective lens and high power objective lens was computed by getting the quotient of ocular and stage micrometer divisions and multiplying it by 10um. The calibration of the two objective lens was compared to indicate the reliability of the computed size of the specimen. The results were recorded in a tabulated form and were discussed. Finally, several research questions pertaining to microscope handling, the microscope itself and the computation for calibration constant were answered.

I.        Introduction

Microorganisms are objects that cannot be seen through the naked eye. (Shamiksa, 2015) But there are certain techniques and instruments used for viewing and measuring them. Micrometry and microscopy are methods that are used in determining the measurements of microorganisms.  Micrometry refers to the practice of measuring microorganisms which uses two micro-scales called ocular micrometer, a circular glass disc that fits into the circular shelf inside the eyepiece of the microscope and a stage micrometer that is clipped to the stage of the microscope. (James & Sherman, 2007) Microscopy is the technical field of using and handling microscopes.

There are two main objectives in this exercise. The first one is to view slides by focusing using both low power objective lens (LPO) and high power objective lens (HPO). The other objective is to measure the size of the cell of Didinium nasutum under the microscope.

II.        Methodology

A well kept compound microscope was used for the exercise of micrometry and microscopy. The  instrument used for calibrating the ocular micrometer was the stage micrometer slide. And an ocular micrometer was used for measuring the size of the specimen, Didinium nasutum .

The microscope was placed on top of the table then a stage micrometer slide was clipped properly at the stage. The microscope was inclined at an angle that will provide enough illumination to the micrometer slide at the stage. Low power objective lens was the first lens to be calibrated. The coarse adjustment knob was adjusted to give a clear view of the slide. The field was adjusted gently so that the zero line of the stage micrometer will be superimposed on the first line of the ocular micrometer. Two superimposed lines were located right next to the first line of the  stage micrometer. The number of space from the first line of the stage micrometer to the first set of superimposed lines was counted. After counting the number of space, the calibration constant was computed by dividing the number of stage micrometer spaces over the number of ocular micrometer spaces and multiplying the quotient by 10 um. The calibration constant  value computed for the low power objective lens was 10 um. The value was written on the data sheet after the computation.

The lens was switched from low power objective lens to high power objective lens. The coarse adjustment knob was adjusted until the slide became clear. The first line of the stage micrometer was superimposed to the first zero line of the ocular micrometer by adjusting the field of view. After making sure that the zero and first line of both micrometers are superimposed and secured, the next pair of superimposed lines was located with  4 ocular micrometer spaces and 1 stage micrometer spaces from the starting lines of both micrometers. The calibration constant of the high power objective lens  were computed by dividing the number of stage micrometer space by the number of ocular micrometer spaces and multiplying its quotient by 10 um. The value of the calibration constant for the high power objective lens, which is 2.5 um, was then recorded into our data sheet.

After calibrating the microscope, the stage micrometer slide was replaced by the Didinium nasutum specimen and the lens was switched into low power objective. The coarse adjustment knob and fine adjustment knob was adjusted to give a clear and fine view of the cells. The field of view was adjusted so that the tip of the cell touches the zero line of the ocular micrometer. The number of divisions that the cell occupies were counted from tip to tip. The length of the cell was computed by multiplying the number of ocular micrometer divisions by the calibration constant of the low power objective lens. We arrived at a value of 140 um and recorded it on our data sheet.

By adjusting the nose piece of the microscope, the low power objective lens was switched into high power objective lens. The coarse adjustment knob and fine adjustment knob were tuned to give a clear and fine view of the cell. By slowly adjusting the field of view, the tip of the cell was made sure that its touching the zero line of the ocular micrometer. The number of divisions that the cell occupies was counted and repeated three times to prevent errors. By multiplying the calibration constant of the high power objective lens to the number of divisions that the cell occupies the length of the cell was calculated. The value, which is  137.5 um, was then recorded into our data sheet.

III.        Results and Discussion

Table 1.1 Computation for Calibration Constant for LPO

For Table 1.1, in all three trials, the number of ocular micrometer that is superimposed to the 1st line of the stage micrometer is 1 thus resulting in a value of 10 um. This value was used as calibration constant in computing the size of the specimen in low power objective lens.

Table 1.2 Computation for Calibration Constant for HPO