Microscopy Text book Chapter 3

Transcription

1 Lab 2 Goals and Objectives: Microscopy lecture: Text Chapter 3 (supplement pg 39) Exercise 1: Brightfield Microscopy Learn how to use the microscope (guide in supplement pg 41) Diopter and interocular adjustments Light adjustments (iris diaphragm) Oil immersion Exercise 5: Microscopic Measurements Calibrate the ocular micrometer (supplement pg 42) Microscope Worksheet (supplement pg 43) Measure the provided specimens Complete the microscope worksheet and turn it in Microscopy Text book Chapter 3 Lab Notes Supplemental Packet page 39 *Microscope Measurement Concept Practice Homework: complete measurement practice sheet in supplemental packet, self check: answers in supplement pg Light Microscope Amy Warenda Czura, Ph.D. 1 SCCC BIO244 Lab 2 Notes

2 Brightfield Phase contrast Darkfield D.I.C. Fluorescence Microscopy D.I.C. Confocal Amy Warenda Czura, Ph.D. 2 SCCC BIO244 Lab 2 Notes

3 Electron Microscopy Scanned-Probe Microscopy DNA Bacterial Surface Protein Positive Staining Negative Staining Gram Stain Acid Fast Stain Amy Warenda Czura, Ph.D. 3 SCCC BIO244 Lab 2 Notes

4 BIO244 Microscopy Capsule Stain Special Stains Students will need:! microscope and cord (sign out number, use all semester)! 1 e slide! 1 stage micrometer! immersion oil: share one bottle per student pair! lens tissue & methanol: share per student pair! lab manual or photocopied pages (Exercises 1 & 5)! laboratory supplemental packet:!guide to Using the Microscope (page 41)!BIO244 Microscope Worksheet (page 43)!Measurements using the Ocular Micrometer (page 42) Exercise 1: Brightfield Microscopy Learn how to use the microscope (guide in supplement pg 41) Diopter and interocular adjustments Light adjustments (iris diaphragm) Oil immersion Exercise 5: Microscopic Measurements Calibrate the ocular micrometer (supplement page 42) Amy Warenda Czura, Ph.D. 4 SCCC BIO244 Lab 2 Notes

5 1.! Make sure left ocular is turned fully counter-clockwise 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) 2.! Start with 10X objective (find it with 4X objective first!) 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) At 100X total magnification they should line up as shown below stage micrometer 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? ocular micrometer 4. Repeat for the 40X objective: 0.1mm = 10 units 1 unit = 0.1mm 10 1 unit = 0.01mm to convert mm to!m, multiply by 1000 When the microscope is set to 100X total micrometer is worth!m Amy Warenda Czura, Ph.D. 5 SCCC BIO244 Lab 2 Notes

6 0.1mm = 40 units 1 unit = 0.1mm 40 1 unit = mm to convert mm to!m, multiply by 1000 When the microscope is set to 400X total micrometer is worth!m When the microscope is set to 100X total micrometer is worth 10!m When the microscope is set to 400X total micrometer is worth 2.5!m When the microscope is set to 1000X total micrometer is worth 1!m 4Xobjective (40X) 4Xobjective (40X) 10Xobjective (100X) 40Xobjective 10Xobjective (100X) 40Xobjective Amy Warenda Czura, Ph.D. 6 SCCC BIO244 Lab 2 Notes

7 4Xobjective (40X) 40X objective 10Xobjective (100X) 100X objective (1000X) 40Xobjective Lab 2 Goals and Objectives: Microscopy lecture: Text Chapter 3 (supplement pg 39) Exercise 1: Brightfield Microscopy Learn how to use the microscope (guide in supplement pg 41) Diopter and interocular adjustments Light adjustments (iris diaphragm) Oil immersion Exercise 5: Microscopic Measurements Calibrate the ocular micrometer (supplement pg 42) Microscope Worksheet (supplement pg 43) Measure the provided specimens Complete the microscope worksheet and turn it in *Microscope Measurement Concept Practice Homework: complete measurement practice sheet in supplemental packet, self check: answers in supplement pg Amy Warenda Czura, Ph.D. 7 SCCC BIO244 Lab 2 Notes