Endocrinology Lab Techniques Flashcards
What is the general purpose of immunohistochemistry techniques?
Visualization of specific cell components
What is the general principle behind the immunohistochemistry method?
Localization of antibody-antigen binding specificity; antibody-antigen interactions are visualized by markers like fluorescent dye, enzymes, radioactive elements, etc.
What is surgical ablation?
The removal of a gland or other organ
What is the removal of the pituitary gland called?
Hypophysectomy
What is the removal of the ovaries called?
Ovariectomy or oophorectomy (spaying)
What is the removal of the testes called?
Orchiectomy (castration)
What is the removal of the adrenal glands called?
Adrenalectomy
What is chemical ablation?
The suppression of endocrine function via chemical means
What is one type of chemical ablation discussed in class?
Alloxan suppresses beta cell function and inhibits the release of insulin
What is deafferentation?
The removal of the nervous stimulation of the tissue
What is a vagotomy?
Disruption of the vagus nerve of gastrin-producing cells of the stomach
What is an in vivo bioassay?
A method that measures the biological response caused by a hormone in a live animal, organ, or tissue
What was an A-Z pregnancy test?
It was an early type of pregnancy test
How did the A-Z pregnancy test work?
The urine of a woman was injected into an immature rat or mouse; if the woman was pregnant, the precence of hCG would cause an estrous reaction in the rat or mouse - the rat or mouse would go into heat despite being developmentally incapable of such
What are three disadvantages of in vivo bioassays?
- A lack of sensitivity - a lot of hormone is usually needed for a response
- Poor reproducibility - wide variability of response in different animals
- High cost and difficulty of use - animals and animal preparations
What are in vitro bioassays?
A technique that employs a cell culture of recommended biological systems to study the effects of a compound under standard conditions
Who developed the radioimmunoassay method?
Rosalyn Yalow
What is radioimmunoassay?
A type of in vitro technique used to measure concentrations of antigens without the need to use a bioassay (the concentration of an unknown labeled antigen is obtained by comparing its inhibitory effect on the binding of radioactively labeled antigen to specific antibody)
What is the principle behind radioimmunoassays?
Competitive binding between a radiolabelled antigen and an unlabelled antigen to a high affinity antibody
What are immunometric assays?
Techniques that use labeled antibodies (not labeled antigens)
What is the principle behind immunometric assays?
Noncompetitive binding
In an immunometric assay the ______________ generated is _____________ to the amount of hormone present.
Color (or other visualization technique)
Directly proportional
What method separates compounds?
Chromatography
What do endocrinologists want to know about a hormone?
Source, structure, biosynthesis, secretion, mechanism of secretion, circulation and metabolism, biological actions and roles, mechanism of action, pathphysiological aspects, and comparitive
Who conducted experiments on male chicks that were castrated, stating, “The testes act upon the blood, and the blood acts upon the whole organism.”
Arnold Berthold
What does CRISPR stand for?
Clusted Regularly Interspaced Short Palindromic Repeats
What is CRISPR?
A genetic engineering tool that uses a CRISPR sequence of DNA and its associated protein (Cas9) to edit the base pairs of a gene
What is the purpose of a Western blot?
To separate and identify specific proteins from a complex mixture extracted from cells
What is the principle behind Western blotting?
It is based on immunochromatography where proteins are separated into polyacrylamide gel according to their molecular weights
What are the three main steps in a Western blot?
Gel electrophoresis
Transfer and incubation
Protein marking via antibodies
What are reporter gene assays used to study?
Gene expression, gene regulation, and/or the structure of regulatory elements that is visualized via a reporter gene protein
What is a luciferase assay?
A reporter gene assay that uses luciferase enzymes as markers of gene expression
What is PCR?
Polymerase chain reaction is a technique that amplifies and frames specific DNA sequences
What is RT-PCR?
RT-PCR creates a complementary DNA fragment from RNA using reverse transcriptase
Why use RT-PCR?
Because RNA is very unstable and difficult to work with
What is Northern blotting used for?
To determine if specific RNA sequences are present within a sample
What is in situ hybridization?
It is a method used to detect the localization of a particular gene
What does RT-PCR provide endocrinologists?
The quantitative measurement of a transcribed gene product from cells and/or tissue
