Lab Exam 1 Study Guide PDF

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Identify the components of a bright field microscope Oculars- lenses that magnify images 10x Right ocular= loose Left ocular= secure Ocular tube- holds oculars & can be adjusted, interpupillary distance (distance b/w eyes) Head-System which helps to send an image to the oculars & your eyes Body-Houses revolving nosepiece & objective lenses Revolving nosepiece-contains 4 objectives at different magnifications Objective lenses-lenses w. mag. of 4x,10x,40x,100x Arm-holds all of the other parts & used to transport microscope Coarse focus knob-on both sides, allows to focus on image Fine focus knob- “fine tunes” on your specimen Base-holds everything in place, used to transport microscope Mechanical stage-where specimen is placed for observation, has a clamp X Stage control knob-knob that moves stage horizontal Y Stage control knob-knob moves stage vertically Condenser system-helps focus light to specimen on a slide Field iris diaphragm-vary diameter of iris diaphragm, limiting the amount of light passing through the condenser system & specimen

Explain what resolution is, and what affects the resolution. Resolution also known as resolving power - the ability to distinguish two very small and closelyspaced objects as separate entities. The resolving power of a microscope is the most important feature of the optical system and influences the ability to distinguish between fine details of a particular specimen. As discussed above, the primary factor in determining resolution is the objective numerical aperture, but resolution is also dependent upon the type of specimen, coherence of illumination, degree of aberration correction, and other factors such as contrast enhancing methodology either in the optical system of the microscope or in the specimen itself. In the final analysis, resolution is directly related to the useful magnification of the microscope and the perception limit of specimen detail.

Give approximately the resolution of our scopes. 0.2 micrometers Explain how to focus, and how to use oil immersion. First focus the lens on the specimen. Then move the 4x in to place in order to put your drop of oil on your cover slip. Once complete, slide the 100x oil immersion lens toward the slide and go past it, then backtrack before clicking into place in order to spread the oil. Once in place, focus and fine tune your lens.

Define parfocal. Parfocal- when 1 lens is in focus, the others have the same focal length and can be rotated into position without making major adjustments to achieve focus Determine magnification of any microscope. 10x (4 or 10 or 40 or 100) Explain how field of view and brightness change with magnification. The light intensity decreases as magnification increases. *As magnification increases, light decreases* Explain how to use the iris diaphragm and when it should be open or closed. move knob on it to the left/right varies diameter of iris diaphragm, limiting the amount of light passing through the condenser system & specimen Determine the field of view of a microscope with a stage micrometer & Give the field of view for your microscope Look at page 14 Estimate the size of a specimen in mm and in µm. Look at page 14

Define: bacillus, cocci, spirochaetes, spirillum, strepto, staphylo. Bacillus-any rods shaped bacterium Cocci-A spherical/oval bacteria Spirochaetes-helical/corkscrew-shaped Spirillum-curved/corkscrew-shaped Strepto-chains of bacterial cells Staphylo- clusters (grapelike) coccobacillus-bacteria that is oval rod

Explain what surfaces or exposures provided the most and the least bacteria for growth. Most- phones, doorknobs, etc Least- gold because it has antibacterial properties

Compare the purpose of blood agar versus other agar media used in our labs so far. Blood agar contains general nutrients and 5% sheep blood. It is useful for cultivating fastidious organisms and for determining the hemolytic capabilities of an organism. Describe and following media: nutrient agar: when agar is added, is becomes a solid medium nutrient broth: is a commonly used liquid complex medium. TSA: Trypticase Soy Agar—TSA has agar so it will be a solid TSB: Trypticase Soy Broth—TSB will be liquid Slant: a solidified medium that has been tilted to provide a greater surface area for growth Deep: a solidified medium that is cooled in an upright position; used when low oxygen levels are required Petri plate: A petri dish containing a growth medium; often used for isolating organism from a Mixed culture: Cultures containing more than one type of microorganism. LB: Luria Broth—a standard medium for maintenance and propagation of E. coli. LB agar: Explain how to place tubes and plates in the incubator. Test tubes: label: name, date, lab #/organism and secure test tube with rubber band Place tube in assigned rack for your table in proper incubator degree Plates: label: name, date, lab #/organism place plate in proper incubator degree

Make a negative stain, or explain how. View lab 7, page 37 Make and explain how to make a smear from a broth place 1 or 2 loopfuls of liquid suspension on the center of the circle spread the suspension into a thin area about the size of a nickel allow smear to air-dry Explain how to make an aseptic transfer from a tube. 1. Incinerate needle / loop 2. Open test tube (1) and wave through heat 3. Wait for needle/loop to cool so you don’t kill the bacteria, then take bacteria from broth or slant 4. wave test tube through heat again and close container 5. Transfer bacteria to the other tube (2) and reincinerate needle Make a simple stain and identify the cells shape and arrangement. Pg. 33 lab book

when smear is all air dry, heat fix by quickly passing the smear over the flame 4 or 5 times place slide on staining tray and cover smear with METHYLENE BLUE DYE for 1 min wash smear with water use bibulous paper and blot dry, don't wipe the slide examine all stained slides under 100x oil immersion lens

Explain and perform a streak plate Pg. 29 lab book

Explain what it means to subculture a bacteria colony. Subculturing describes the transfer of microbes from one growth medium container, such as broth or agar, to another, and allowing the microbes to grow.

Perform and list all the steps and their timing of a Gram stain, note the results of a positive and negative stain 1. Put DI water on needle 1. Add drop on the slide 2. Aseptically get the bacteria on the needle (incinerate needle, heat test tube, once needle cools stick into test tube and get bacteria on the needle, heat test tube and close) 3. Transfer bacteria on the needle to the slide where the drop of DI water is. 4. Wait til it air dries, then heat fix (wave through burner 3-4 times) 5. Flood smear with crystal violet for 30 seconds 6. Wash with DI water to remove excess dye 7. Flood with iodine for 1 minute and wash with DI water to remove excess stain 8. Decolorize with alcohol wash (hold at angle and only 3-5 seconds then immediately wash with DI water) 9. Counterstain with safranin for 1 minute, then wash off excess dye with DI water 10. Blot with bibulous paper 11. Place slide on microscope, bring to focus and add oil for oil immersion

Voice MemoBroth streak plate--label it correctly. Put it in 30 degrees -looking for: did you label it, if there is contamination, do it any style you want Slant gram stain- is gram positive or negative? What cell shape is it? cocci? Bacillus? (probably going to either coccus or bacillus). Tell the arrangement-- is it Stepto? Staphylo? Or single?

Gram positive- purple Gram negative- Pink Hanging drop- slant (di water) or broth. Be able to tell if its motile or not. Short answer: How do you do gram stain, how you prepare a smear, negative stain vs a normal stain, acid vs basic dyes, how to maintain microscope & how to use it (know all parts and what they do). Measurement-tell how big it is and how you measure it. Estimate. FOV. Fill in: thing to measure, motility, gram stain, streak plate...