MIBO Lab Midterm Study Gu

MIBO Lab Midterm Study Gu - MIBO Lab Final Study Guide...

Info iconThis preview shows pages 1–4. Sign up to view the full content.

View Full Document Right Arrow Icon
MIBO Lab Final Study Guide Activity 1: Lab Saftey Activity 3: Brightfield Microscopy Parts and functions of microscope Total magnification = objective lens x ocular lens Oil Immersion o Start at 40x, then turn halfway to 100x o Add drop of immersion oil to slide o Change to 100x lens o Use fine focus to adjust o Clean with isopropyl alcohol and lens paper Focusing a microscope Activity 4: Smear Prep and Simple Staining Types of dye o Acidic dye (anionic) o Basic dye (cationic) Components of dyes o Auxochrome : binds stain to oppositely charged cell components o Chromophore : colored group attached to auxochrome Simple stain o Single dye applied to smear Positive simple stain : colors cells Negative simple stain : colors background o Observations: Morphology ( coccus, bacillus, spirillum ) Cellular arrangements ( diplo, strepto, staphylo, tetrad 4 , sarcina 8) Smear Prep from Plate w/ Simple Stain o Loopful water on slide o Aseptic transfer of culture, spread it out o Air dry and heat fix o Dye for 3 minutes (crystal violet or methylene blue) o Rinse, blot, air dry Smear Prep from Broth w/ Simple Stain o Aseptic transfer culture, spread it out o Repeat 2-3 loopfuls o Air dry and heat fix o Dye for 3 minutes (crystal violet or methylene blue) o Rinse, blot, air dry Activity 2: Handling and Examining Cultures
Background image of page 1

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Broth Aseptic Transfer o Make sure bubble doesn’t pop Plate Aseptic Transfer o Keep lid closed as much as possible o Incubate cultures in inverted position so that water does not condense on surface of agar Activity 5: Gram Stain Primary dye : Crystal violet o Stains all organisms Mordant : Gram’s Iodine o Strengthens the union b/t the dye and its substrate Decolorizer : Ethyl alcohol o Removes primary stains from cells Secondary dye : Safranin o Stains all cells to add colors to decolorized cells Procedure o Smear, air dry, heat fix o Flood with crystal violet for 60 seconds (primary dye) o Rinse w/ water o Flood with Gram’s Iodine for 60 seconds (mordant) o Rinse w/ water o Decolorize with a few drops of ethyl alcohol o Rinse w/ water o Flood with safranin for 30 seconds o Rinse w/ water o Blot dry Results o Gram + stain purple Have thick layer of peptidoglycan Pores in cell wall are closed and retain crystal violet o Gram – stain pink Higher lipid content in cell wall Lipids dissolved by alcohol, crystal violet leaks out Pitfalls o Decolorization is critical Too much: gram + appear – Too little: gram – appear + o Culture must be less than 24 hours old Older cultures’ cell wall will break down and Gram + can’t retain dye Activity 6: Acid Fast Stain and Endospore Stain
Background image of page 2
Acid Fast o Primary stain : carbolfuchsin o Decolorizer : Acid-alcohol o Secondary stain : Methylene blue Procedure o Smear, air dry, heat fix o Flood with carbolfuchsin o Steam over boiling water for 5 minutes, adding more dye as necessary o Cool, rinse w/ water
Background image of page 3

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Image of page 4
This is the end of the preview. Sign up to access the rest of the document.

This note was uploaded on 06/21/2011 for the course MIBO 2500 taught by Professor Walker during the Spring '08 term at UGA.

Page1 / 12

MIBO Lab Midterm Study Gu - MIBO Lab Final Study Guide...

This preview shows document pages 1 - 4. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online