## 1) contrast to see. Light microscope magnification

1)   Explain the general concept ofhow a brightfield microscope works.  Whatare the major limitations? (1pt) Brightfield microscope workingprocedure; In microscopy afterlocating a sample on the stage, light comes from the light source . There is alens which is named as condenser aimed the sample.  The condenser has a diaphragm which hasaperture.

this diaphragm is for controlling and focusing the light on thesample. Light passes through the sample. this light is collected by anobjective lens. This lens is on the stage. The objective helps the light morevisible.

The user can see the sample from oracular lens or eyepiece. Thecontrast, which isabsorbed by stains, pigmentation, or dense areas by light, allowsyou to see the sample. Major limitations are; Somebiological samples have low contrast to see. Light microscope magnification isaround 1300X.

There are bigger magnifications but if you increase it, theimage’s clarity will decrease. Naturally colorless and transparent samples arevisible. They should be stained before using the brightfield microscopy.  2)   What is the RefractiveIndex?  What is the working distance? (0.5pt) Refractive index is the ratio of the velocity of light ina vacuum to its velocity in a specified medium. For calculation theworking distance, Numerical aperture is used. Numericalaperture is basically for how much good detail you can see.

The numericalaperture formula is Numerical Aperture (NA) = ? • sin(?) a = one-half of the objective’s opening angle ? = refractiveindex of the immersion medium                        Refractive index is the limitingfactor to achieve numerical apertures more than 1.0. So, to get biggernumerical apertures, the refractive index must be increased.  3)   What is the totalmagnification when using the 4X, 10X, 40X, and 100X objectives? (0.

5pt) Total magnification is calculated by multiply the objectives (4X, 10X, 40X,and 100X) and power of eyepiece (10X) TotalMagnification;(4X)* (10X) = 40X(10X)* (10X) = 100X(40X)* (10X) = 400X(100X)* (10X) = 1000X 4)   Bacterial species are rarelyidentified by colony morphology alone. Explain why this is usually the case. Give one or more example(s) ofother techniques that may aid in identifying an unknown microorganism.

(1pt)Bacterial morphology deals with size, shape, and chromogenesis, opacity, elevatin,surface. margin generally, these criteria may be enough to identify thespecies. Also Aggregation andbiochemical testing are techniques to identify the microorganisms.  5)   Define differential stain.

Howis this technique different from a simple stain? (0.5pt) Differentialstain works with chemical as simple stain. In this technique colors areimportant as simple stain. But there are more than 3 chemicals. The differentialstain does not give the same color to the bacteria. Differential stain is foridentify the cell types, their structure basis on their different colors.

6)   Compare and contrast cellwalls of G+ and G- bacteria. (0.5pt) ·      Gram+ bacteria has thick (multilayered) peptidoglycan layer. Gram- bacteriahas thin (single-layered) peptidoglycanlayer.·      Gram+ bacteria has not a outer membrane. Gram- bacteria has a outermembrane.·      Two of them has a cytoplasmic membrane. ·      Gram+ bacteria has low lipid andlipoprotein content.

Gram- bacteria has high lipid and lipoprotein content. ·      Gram+ bacteria is More susceptible toantibiotics.Gram- bacteria is More resistant to antibiotics.

7)   Make a table with the color ofE. Coli and S. Epidermidis after each step you completed during your Gramstains in lab (Crystal Violet, Iodine, Alcohol, Safranin). (1pts)  E. Coli S. Epidermidis Primary Stain Crystal Violet Purple Purple Mordant Iodine Purple Purple Decolorizer Alcohol Colorless Purple Counterstain Safranin Pink Purple    “BrightfieldMicroscopy – Uses & Advancements; Microscope Reviews; Pros and Cons.” MicroscopeMaster, www.microscopemaster.

com/brightfield-microscopy.html. “Bright-Fieldmicroscopy.” Wikipedia, WikimediaFoundation, 19 Jan.

2018, en.wikipedia.org/wiki/Bright-field_microscopy. “Refractiveindex.” Wikipedia, WikimediaFoundation, 29 Jan. 2018, en.wikipedia.org/wiki/Refractive_index.

“Propertiesof Microscope Objectives.” Nikon’s MicroscopyU,www.microscopyu.com/microscopy-basics/properties-of-microscope-objectives.

http://zeiss-campus.magnet.fsu.edu/articles/basics/resolution.html “Numericalaperture.” Wikipedia, WikimediaFoundation, 31 Dec. 2017, en.wikipedia.

org/wiki/Numerical_aperture. “UnderstandingYour Objective.” Thermo Fisher Scientific, www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-analysis-learning-center/molecular-probes-school-of-fluorescence/fundamentals-of-fluorescence-microscopy/understanding-your-objective.html.  http://www.cas.

miamioh.edu/mbiws/microscopes/Magnification.html Libretexts.”8: Bacterial Colony Morphology.” Biology LibreTexts, Libretexts, 2 Jan. 2018,bio.libretexts.

org/Labs/Microbiology_Labs_I/08%3A_Bacterial_Colony_Morphology. “Chegg.com.” Definition ofBacterial Morphology | Chegg.

Com, www.chegg.com/homework-help/definitions/bacterial-morphology-14. “Bacterialcellular morphologies.” Wikipedia, Wikimedia Foundation, 22 Jan.

2018,en.wikipedia.org/wiki/Bacterial_cellular_morphologies. http://www.nios.ac.

in/media/documents/dmlt/Microbiology/Lesson-11.pdf drillmaster2001.”What is the difference between a differential and a simple stainDifferential.” Simple stainDifferential,www.coursehero.

com/file/p18b4vm/What-is-the-difference-between-a-differential-and-a-simple-stain-Differential/. “Gram-Positivevs Gram-Negative Bacteria.” Gram-Positivevs Gram-Negative Bacteria – Difference and Comparison | Diffen, www.diffen.

com/difference/Gram-negative_Bacteria_vs_Gram-positive_Bacteria.