Bright Field Microscope, Preparing Tissue Samples

What is the Bright field microscope and How does it work.

Bright field microscopy is the basic type of microscopy.

Biological specimens are viewed by contrast which works through the absorption of stain by

a specimen(Microscopemaster.com, 2014). The specimen, non living, ( cells and tissues)

may not be naturally highly absorbing, so the specimen is stained so that cellular features

can be identified (Berkeley Physics, 2014). The bright field microscope works by aiming a

light towards the collector lens, here the condenser focuses the amount of light specified

through the specimen, and with staining and other techniques the specimen can be viewed

clearly through magnification. (Microscopy U – The Source for Microscopy Education, 2014)

Steps in preparing a tissue section, ready for mounting on a microscope slide and ready for staining.

• Fix your specimen, it preserves the tissue (Cuyahoga County Medical Examiner,

2014) and keeps it attached to the slide during the staining process. (Rolls and

Chapman, 2014)

• Dehydrate it (it should be washed after fixation)

• Clearing is the removal of the dehydrating agent

• Embedding: The tissue will have it’s liquid replace with Paraffin wax, making it hard

enough to be stained and used. (Man Anatomy, 2014).

• Sectioning: Tissue is sliced thinly

• Now the specimen is able to be mounted and stained.

The Haematoxylin and Eosin staining method, its purpose and the results.

Method:

• Dip blood smear in Fixer five times for one second each time. Drain off remaining

liquid

• Repeat when dipping in Red Stain

• Repeat when dipping in Blue Stain

• Wash stain with distilled water and allow to dry (VetLogicVideo, 2007)

It is used to provide Histological examination of tissue sections. Because it is charged-Based

Hematoxylin, the base will stain Basophilic parts blue (Histology University of Leeds. 2014)

(these parts are positive so they will attract a negative dye (Feinberg Northwestern

University, 2014)) and Eosin stains acidophilic parts (basic pH and are mainly Cytoplasmic,

such as protein products) pink (Fischer and Jacobson et al., 2008).

References:

Berkeley Physics PHYS250. 2014. Berkeley PHYS250 Lecture 2 Microscopes.

http://physics.berkeley.edu/research/yildiz/Teaching/PHYS250/Lecture_PDFs/Lecture_2.pdf

Viewed on the 10th of February 2014

Cuyahoga County Medical Examiner. 2014. The Steps for Preparation of Slides – Cuyahoga

County Medical Examiner

http://medicalexaminer.cuyahogacounty.us/en-US/Steps.aspx Viewed on the 11th of

February 2014

Feinberg Northwestern University. 2014. The Science and Application of Hematoxylin and

EosinStaining.

http://www.feinberg.northwestern.edu/research/docs/cores/mhpl/HandE_troubleshooting.pdf

Viewed on the 10th of February 2014

Fischer, A., Jacobson, K., Rose, J. and Zeller, R. 2008. Hematoxylin and Eosin Staining of

Tissue and Cell Sections. Cold Spring Harbor Protocols, 2008 (5), pg 4986.

Can be Viewed at : http://www.ncbi.nlm.nih.gov/pubmed/21356829 Accessed on the 10th of

February 2014

Gill, G. W. 2014. Chapter 13 H&E Staining: Oversight and Insights.

http://www.dako.com/08066_12may10_webchapter13.pdf Viewed on the 10th of February

2014

Histology University of Leeds. 2014. Histology Guide | What Is Histology.

http://histology.leeds.ac.uk/what-is-histology/H_and_E.php Viewed on the 11th of February

2014

Man Anatomy. 2014. Preparation of tissue for microscopy.

http://www.mananatomy.com/histology/preparation-tissue-microscopy Viewed on the 11th of

February 2014

Microscopemaster.com. 2014. Brightfield Microscopy – Uses and

Advancements; Advantages and Disadvantages.

http://www.microscopemaster.com/brightfield-microscopy.html Viewed on the 10th of

February 2014

Microscopy U – The Source for Microscopy Education. 2014. Nikon MicroscopyU |

Fundamentals of Digital Imaging.

http://www.microscopyu.com/articles/digitalimaging/digitalintro.html Viewed on the 11th of

February 2014

Rolls, G. 2012. Performing a Hematoxylin and Eosin Stain – a Step by Step Guide.

http://www.leicabiosystems.com/pathologyleaders/performing-a-hematoxylin-and-eosinstain-

a-step-by-step-guide/ Viewed on the 10th of February 2014

Rolls, G., Chapman, C., Rasanen, M. and Stephen Peters, D. 2014. Histology Sample

Preparation. http://www.leicabiosystems.com/pathologyleaders/topics/histology-samplepreparation/

Viewed on the 10th of February.

VetLogicVideo, 2007, Making a blood smear, http://www.vetlogic.uk.co Viewed on the 7th of

February 2014

VetLogicVideo, 2007, Staining a blood smear http://www.vetlogic.uk.co Viewed on the 7th of

February 2014

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