BIO244 Lab 1 MicroscopyStudents will need:

• microscope and cord (sign out number, use all semester)• 2 sterile agar plates, stack and tape to keep lids shut• 4 sterile swabs• 1 vial sterile water• 1 “e” slide• 1 stage micrometer• immersion oil: share one bottle per student pair• lens tissue & methanol: share per student pair• laboratory supplemental packet• lab manual or photocopied pages• BIO244 Microscope Worksheet• Chapter 3 Notes outline

MicroscopyText book Chapter 3

Lab Notes

Light Microscope

http://www.microscopy-uk.org.uk/index.html?http://www.microscopy-uk.org.uk/intro/illu/illu.html

Brightfield Darkfield

Phase contrast D.I.C.

Fluorescence Microscopy

D.I.C.Confocal

Electron Microscopy

DNA Bacterial Surface Protein

Scanned-Probe Microscopy

Positive Staining Negative Staining

Gram Stain

Acid Fast Stain

Capsule Stain

Special Stains

Lab 1 Goals and Objectives:

Microscopy lecture: Text Chapter 3

Exercise 1: Brightfield MicroscopyLearn how to use the microscope (guide in suppl. packet p.51)

Diopter and interocular adjustmentsLight adjustments (iris diaphragm)Oil immersion

Exercise 5: Microscopic MeasurementsCalibrate the ocular micrometer (suppl. packet p.52)

Microscope WorksheetMeasure the provided specimensComplete the microscope worksheet and turn it in

Microscope Measurement Concept Practice Homework: complete measurement practice sheet in supplemental packet, self check: answers in packet (p. 53-54)

Exercise 7: Ubiquity of Bacteria (Replacement activity in packet p. 55)Homework: “Testing Household Surface” handout, Take home plates, swabs, and sterile waterREAD DIRECTIONS CAREFULLY! then do experiment

Exercise 7: Ubiquity of Bacteria (Replacement activity in packet p. 55)Homework: “Testing Household Surface” handout, Take home plates, swabs, and sterile waterREAD DIRECTIONS CAREFULLY! then do experiment

Exercise 1: Brightfield MicroscopyLearn how to use the microscope (guide in suppl. packet p.51)

Diopter and interocular adjustmentsLight adjustments (iris diaphragm)Oil immersion

Exercise 5: Microscopic MeasurementsCalibrate the ocular micrometer (suppl. packet p.52)

1. Make sure left ocular is turned fully counter-clockwise

2. Start with 10X objective

3. Get stage micrometer (2mm ruler on slide) in focus: you will see the ocular micrometer with labels in multiples of 10 (0, 10, 20, 30) and the stage micrometer with labels in multiples of 0.1 (0.0, 0.1, 0.2)

stage micrometer

ocular micrometer

3. Line up the zero lines of the two micrometers (you can turn the top of the ocular to position the ocular micrometer and the mechanical stage controls to move the stage micrometer)

At 100X total magnification they should line up as shown below

“0.1” on stage micrometer is 0.1mm on that actual 2mm ruler. This distance corresponds to 10 “units” (spaces between lines) on the ocular micrometer (0-10): the 0.1 line of the stage ruler lines up with the 10 line on the ocular one. How long in millimeters is one unit on the ocular micrometer with the 10X objective in place?

0.1mm = 10 “units”

1 unit = 0.1mm ÷ 10

1 unit = 0.01mm

to convert mm to µm, multiply by 1000

When the microscope is set to 100X total magnification, one unit on the ocular micrometer is worth ________µm

4. Repeat for the 40X objective:

0.1mm = 40 “units”

1 unit = 0.1mm ÷ 40

1 unit = 0.0025mm

to convert mm to µm, multiply by 1000

When the microscope is set to 400X total magnification, one unit on the ocular micrometer is worth ________µm

When the microscope is set to 100X total magnification, one unit on the ocular micrometer is worth ____10____µm

When the microscope is set to 400X total magnification, one unit on the ocular micrometer is worth ____2.5____µm

When the microscope is set to 1000X total magnification, one unit on the ocular micrometer is worth ____1____µm

DinoflagellateDinoflagellate 4Xobjective(40X)

10Xobjective(100X)

40Xobjective(400X)

Czura 2005

Lilium PollenLilium Pollen 4Xobjective(40X)

10Xobjective(100X)

40Xobjective(400X)

Czura 2005

Frog BloodFrog Blood 4Xobjective(40X)

10Xobjective(100X)

40Xobjective(400X)

Czura 2005

100X objective(1000X)

40X objective(400X)

Streptococcus/Enterococcus faecalisStreptococcus/Enterococcus faecalis

Czura 2005

DinoflagellateDinoflagellate

Frog BloodFrog Blood

Lilium PollenLilium Pollen

Streptococcus faecalisStreptococcus faecalis

Lab 1 Goals and Objectives:

Microscopy lecture: Text Chapter 3

Exercise 1: Brightfield MicroscopyLearn how to use the microscope (guide in suppl. packet p.51)

Diopter and interocular adjustmentsLight adjustments (iris diaphragm)Oil immersion

Exercise 5: Microscopic MeasurementsCalibrate the ocular micrometer (suppl. packet p.52)

Microscope WorksheetMeasure the provided specimensComplete the microscope worksheet and turn it in

Microscope Measurement Concept Practice Homework: complete measurement practice sheet in supplemental packet, self check: answers in packet (p. 53-54)

Exercise 7: Ubiquity of Bacteria (Replacement activity in packet p. 55)Homework: “Testing Household Surface” handout, Take home plates, swabs, and sterile waterREAD DIRECTIONS CAREFULLY! then do experiment