Eli Flashcards
Order of Leukocyte Frequency
- Neutrophils
- Lymphocytes
- Monocytes
- Eosinophils
- Basophils
Never Let Monkeys Eat Burritos
Neutrophil
Appearance & Function
Granulocyte
Multiple nuclei & light granules
Phagocytosis
Eosinophil
Appearance & Function
Granulocyte
Red/ orange granules
Helminth defense
Basophil
Appearance & Function
Granulocyte
Dark granules
Allergic reactions
Monocyte
Appearance & Function
Agranulocyte
One bean shaped nucleus
Phagocytosis
Lymphocyte
Appearance & Function
Agranulocytes
Small and dark (same size as RBCs)
Specific immunity
Serology
use of antigen-antibody interactions to diagnose disease
Antigen
component of pathogen that stimulates the
immune system
Antibody
proteins made by B-cells that bind
to the antigen
ELISA
Enzyme linked immnosorbent assay
Direct ELISA
Antigen (pathogen): Patient sample
Antibody (patient serum): Produced in the lab
Indirect ELISA
Antigen (pathogen): Produced in the lab
Antibody (patient serum): Patient sample
ELISA Steps
- Antigen is coated onto the test plate
- Primary antibody (from patient serum) is added
- Secondary antibody (anti-human; has an enzyme tagged on it) is added
- Substrate for enzyme is added
- (+) = color change
If a patient doesn’t have the primary antibody, will a color change occur?
No
If a positive color change occurs, does that confirm that the patient has disease?
No. A positive ELISA only indicates that the patient has antibodies against a particular pathogen.
Lysozyme
enzyme that destroys bacterial peptidoglycan by
targeting the NAG-NAM connections
• Part of the innate immune system
• Found in tears, saliva, and sweat
What media do you use for Lysozyme and Tears?
What was the process like?
NA Plates
in 6 different dilutions like pizza slices
What media do you use for saliva?
What was the process like?
Pathogen?
Blood agar plate
4 quadrant swabing a lawn on one
then streaking w/loop on the others
creating isolation
Pathogen: Streptococcus pygoenes
What media do you use for microbes of the skin?
What was the process like?
MSA Plate
4 quadrant swabing a lawn on one
then streaking w/loop on the others
creating isolation
Pathogen: Staphylococcus aureus microbes of the skin
Does lysozyme affect G+ bacteria, G- bacteria, or both?
G+ bacteria
Why do you think lysozymes are more effective against
G+ bacteria?
Lysozymes destroy peptidoglycan cell wall
Blood agar plate
Selective: nothing
Differential: hemolysis
Gamma = no hemolysis Alpha = partial hemolysis Beta = complete hemolysis
Pathogen: Streptococcus pyogenes = Beta hemolytic
Mannitol salt agar (MSA) plate
Selective: halophiles (growth)
Differential: mannitol fermentation (positive = yellow)
Pathogen: Staphylococcus aureus = positive for mannitol
fermentation
Limitations of the Elisa test for patient diagnosis?
- a positive result only confirms there is an antibody present. but it does not mean the patient is currently ill.
- People who are poor producers of antibodiesor who have an interfering substance may get a false positive because the antibody amount is too low and can go undetected.
What are the main reagents added to the ELISA test.
- ANTIGEN
- PRIMARY ANTIBODY
- SECONDARY ANTIBODY
- SUBSTRATE
ANTIGEN
Component of a pathogen that will stimulate the immune system. Coats the wells.
PRIMARY ANTIBODY
Patients serum antibodies. If antibodies against the antigen are present then they will bind to the antigen that is at the bottom of the wells.
SECONDARY ANTIBODY
Animal antibody solution. Antibody against human antibodies which is covalently conjugated to an enzyme.
SUBSTRATE
Colorgenic enzyme substrate solution. Interacts with the enzyme on the second antibody which generates a visible color.
What is the purpose of having a negative control in ELISA?
Incase enzymes they stick to the plastic sides and create a false positive.
Why is is necessary to rinse sample wells between reagent steps?
Rinsing will remove the antibodies that did not react with the antigen. And you can get a false positive if not rinsed.
What should you observe as a positive result for the ELISA test?
What does this tell you about the patient?
A positive result will result in a color which tells us the patient has antibodies
Initial HIV testing is performed via an ELISA
Which immunologic method is used as the verification test for a positive ELISA HIV result?
Western blot test
Separates proteins by electrophoresis and uses antibodies to detect proteins.
