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1.3um, a 10x coular lens, and a 100x oil immersion lens, would you be
able to discern two objects separated by 3um? 0.3um? 300nm? Why would,
or would not you be able to do this with thge present microscope?
Visit http://www.meijitechno.co.uk
Sorry I did not give you enough stuff. This is a micro class and it is blood and bacteria, this is a regular compound light microscope with resolving power of 2um , and yes disern means to separte three parts of the bacteria hope this help. smmity3360
When you say "discern two objects separated by 3um? 0.3um? 300nm? Why would,
or would not you be able to do this with thge present microscope?" do you mean two objects in the field that are separated by the distance of 3 um? OR do you want to know the resolving power of a compound microscope?
What kinds of objects are you viewing?
Bacteria?
Blood cells?
Tissue?
What kind of compund 'scope are you talking about? For a clinical laboratory? A crime laboratory? You would need an electron microscope to see anything smaller than bacteria, or anything much smaller than 1um. Also, you would not use the 10x occular and the 100x oil immersion lens at the same time!
Thank you!
Sincerely,
crabcakes
I?m still not completely sure what you are asking, but I?m going to tackle what I think you mean. Please ask for an Answer Clarification, before rating, if my answer is not what you were hoping for.
?The typical compound light microscope is capable of increasing our ability to see detail by 1000 times so that objects as small as 0.1 micrometer (um) or 100 nanometers (nm) can be seen.? http://abacus.bates.edu/~ganderso/biology/resources/microscopy.html
With a compound microscope, using an oil immersion lens, you would be able to see things as small as .1 um (micrometer),and discern things that are 0.1 um apart.
With a 10x compound light microscope, such as used in clinical laboratories (Usually Zeiss or Nikon), you would typically use a: 10x lens for scanning a slide for the presence of bacteria, tissue, Fern tests, a quick WBC estimate before doing a manual differential, etc.
Using the 40x lens (high dry) you can enumerate cells, scan for abnormal WBC and RBC, RBC morphology, perform a manual platelet count, manual WBC count, sperm counts, etc., do a manual diff (if you are experienced), do a urine microscopic, etc.
With the 100x oil immersion lens you perform platelet estimates, morphology (WBC) and determine bacterial species (Gram stain), Tzanck prep, etc. 100X oil immersion if for fine detail. You can see granules in stained eosinophils and basophils. You can see some WBC cell division (mitosis) in immature WBC, see iron deposits in siderocytes, see malarial parasites in an RBC, and even an Auer rod in a myeloblast, indicating myelocytic leukemia. Because good microscope lenses are parfocal, you can quickly switch from 10x to 40x to 100x without losing your field of view.
Auer Rod
http://udel.edu/~rmaser/hematology/wbc/AUERROD.JPG
Here is a picture of several WBCs, using 100x oil
http://www-medlib.med.utah.edu/WebPath/HEMEHTML/HEME100.html
?Determining Field-of-View Diameter
You may wish to estimate the size of the specimens (e.g., cells) you will see in lab. The best way to do this is with an ocular micrometer, a precision ocular lens insert that has a ruler etched into glass. The monocular scopes we use in the introductory courses are not so equipped, so we will use an alternative method based upon knowing the field-of-view diameter for your particular microscope. To do this, you must determine: ·the approximate diameter of your low magnification field-of-view for your particular microscope. ·the total magnification for each of your other objective lenses.
Knowing this for each objective lens, you can compare the size of the specimen against the known field diameter and make a reasonable esimate of size. This technique works for any microscope. 1. Obtain a slide scale and position it on your scope. A transparent metric ruler will work as well. 2. Bring it into focus using the 10x objective (100x total). The scale bars are increments of 1mm as shown in the figure below. Thus, a black bar = 0.5mm as does a space. 3. Move the slide such that the edge of an outside black bar is just tangent to the lighted field (see point "A" above). 4. Starting at that edge, estimate how many bars and spaces it takes to cross the field-of-view. You will probably have to estimate the last fraction of a space or bar. In the figure above, it is approximately 1.8mm wide. 5. Record your scope's ID number and field diameter at 100x in your lab notebook for future reference. 6. Next, calculate the field width at 430x total magnification using the following formula (we refer to the 100x mag as "low power" and 430x as "high power"): (low power mag/ high power mag) x low power field diameter (in mm)
For the example above,
(100 / 430) x 1.8 mm = 0.418 mm = 418 um (micrometers)
Note that the field diameter at high power is proportional to the ratio of the low to high power objectives. That is, as you increase magnification, the actual field of view becomes proportionally smaller.? http://abacus.bates.edu/~ganderso/biology/resources/microscopy.html
Bacteria are about 2 micrometers in size. A healthy RBC (Red blood cell) is about 7 micrometers across. Platelets are 1-4 micrometers in size. (It is unknown how big knifelets and forklets are - sorry, I couldn?t resist some clinical humor) Using a compound light microscope, you would be able to determine if bacteria had a capsule(faintly), if it were gram negative or gram positive, some flagella, and the size of bacteria, but you would not be able to see ribosomes, DNA, or much separation of cell wall and cytoplasm. For that you'd need an electron microscope.
Bacteria
http://www.disknet.com/indiana_biolab/b003.htm
RBC
http://www.wadsworth.org/chemheme/heme/microscope/rbc.htm
Platelets
http://www.wadsworth.org/chemheme/heme/microscope/platelets.htm
http://www.dmacc.cc.ia.us/instructors/scottie.htm
See all the parts of a compound microscope here:
http://www.cas.muohio.edu/~mbi-ws/microscopes/microscopeparts.html
This site gives you an idea of the size of microscopic things
http://www.cellsalive.com/howbig.htm
More about compound microscopes
http://www.mansfield.ohio-state.edu/~sabedon/biol4030.htm
As you can see, of course you can discern bacteria that are 3 micrometers apart.
This page has a rather blurry picture of bacteria in a high dry(400x) vs. oil immersion fields (100x). http://biology.clc.uc.edu/fankhauser/Labs/Microscope/Oil_Immersion/07_compare_400x_1000x.jpg
?Use an oil immersion lens when you have a fixed (dead - not moving) specimen that is no thicker than a few micrometers. Even then, use it only when the structures you wish to view are quite small - one or two micrometers in dimension. Oil immersion is essential for viewing individual bacteria or details of the striations of skeletal muscle. It is nearly impossible to view living, motile protists at a magnification of 1000x, except for the very smallest and slowest.?
?Note that you can see some detail at 400x, but the shapes and colors of the bacteria are somewhat distorted. Move the 400x lens out of the way, place a drop of immersion oil directly on the smear where the objective was, and swing the oil immersion lens in place. Move the fine focus up and down slightly to ensure that the lens is in contact with the oil. Now watch the end of the objective and bring it as close to the slide surface as you can without touching it. Note in which direction you must focus to move the objective away from the slide, look in the eyepieces, and slowly rotate the fine focus control until the image is focused. Most bacterial species are rod-shaped or round (cocci), although some are curved, spiral-shaped, or irregularly shaped. The gram stain leaves some cell types pink (Gram negative) and others dark blue (Gram positive) depending on cell wall characteristics. Gram stain results are a major criterion for identification of species. Note how much more clear the image is at 1000x with oil than it was at 400x without oil.? http://www.ruf.rice.edu/~bioslabs/methods/microscopy/oilimm.html
Here you can see the difference in magnification between electron and light microscopy. e.coli as seen in an electron microscope
http://www.microbelibrary.org/images/mketterer/Images/mketng.jpg
e.coli using a compound light microscope, 100x oil immersion
http://www.courses.ahc.umn.edu/medical-school/IDis/Images/E.coli.gif
This page contrasts electron and light microscopy
http://www.bact.wisc.edu/Bact330/lecturenf
Principals of Light Microscopy
http://www.life.umd.edu/CBMG/faculty/wolniak/wolniakmicro.html
Using a microscope is one of the most enjoyable aspects of laboratory science. I found it exhilarating to be the one to find the leukemic cell, the malarial parasite, or the two-headed sperm. (When you get to the point where you can identify trichomonas in urine or a wet prep, try this trick: add a drop of KOH under the cover glass, and watch through the oculars as the KOH floods the field. Within a few seconds, the trichomonads will swell and burst! (Sort of like watching an old Space Invaders game!)
Hope this helps you increase your knowledge of microscopy!
Regards
crabcakes
Oh, don't forget to wipe off the oil immediately after using an oil immersion lens!
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