December 2012 - MicrobeHunter.com

Microscope calibration techniquesMEASURINGA comparison of a cheap versus a more expensive method.Roland LutsTwo years ago, I performed a calibrationof my old Reichert compoundmicroscope (from aroundthe year 1960). Lucky for me, this microscopewas already equipped with anocular micrometer. Having not enoughmoney at that time to buy a stage micrometer,I searched the Internet forother means of calibration and found aneasy way, good enough for my purpose:to be able to measure the length of microscopicspecimen in the range of 10 to500 micrometers with reasonable accuracy.See the Internet:www.microscopyuk.org.uk/mag/artmay00/measure.htmlThis is an article of Jean-Marie Cavanihacwith title “Introduction to quantitativemicroscopy – Measuringdiatoms with a ruler”. A part of thisprocedure inspired me to do my calibration.The cheap methodThis cheap method consists in measuringthe known thickness of a coverslip(in terms of total number ofdivisions) with the aid of my ocularmicrometer for all 4 of my objectives(X4, X10, X40, X100). Most coverslipthicknesses are stated to be between 150and 170 micrometers so I assumed minewas 160 micrometers thick as the meanvalue. In a series of photographs I showyou how I mounted a piece (splinter) ofcoverslip on an object glass with chewinggum or other means and what I haveseen through the ocular (figure 1).Painting the coverslip with a markergave even better contrasting results.1Figure 1: Coloured broken coverslipmounted on object glassFigure 2: Coloured broken coverslipmounted on object glass undermicroscope objective2Objective Calibration with cover glass Calibration with stage micrometer1 division on the ocular micrometer 1 division on the ocular micrometercorresponds to:corresponds to:X4 40 micrometers 38.5 micrometersX10 16 micrometers 14.4 micrometersX40 4 micrometers 3.75 micrometersX100 1.6 micrometers 1.50 micrometersMicrobeHunter Microscopy Magazine - December 2012 - 27