MIC: Lecture Ch 17 - Immune and Nucleic Acid Testing

what is immune testing

serology-study and diagnostic use of antigen-antibody interactions in blood serum

what are the 2 general diagnostic processes of immune testing

use known antibodies (from Biotech) to detect antigens associated with an infectious agent
- use known antigens (from Biotech) to detect specific antibodies in a patient’s blood determining exposure to specific pathogen

what is the immune testing choice based

- suspended diagnosis
- cost to perform the test
- speed with which a result can be obtained

what are the types of serologic test

- agglutination test
- labeled antibody test: ELISA and Westen blot
- nucleic acid test (NAT)

what is an agglutination test

cross-linking of antibodies with particulate antigens resulting in clumping of insoluble particles
→ this test takes a couple of minutes to perform and results are easy to interpret with unaided eye

what is the agglutination test used to diagnose

- bacteria: staphyloccus and streptococcus ssp
- viral: hepatitis B
- toxin: tetanus
- drugs
- ABO blood typing

what is the labeled antibody test ELISA and what does it detect

Enzyme-Linked ImmunoSorbent Assay → used to detect either antigens or antibodies from a sample - it can quantify amounts of antigen or antibody

what type of antibody does the ELIZA use

2 degree antibody molecule linked to a label enzyme that allows for substrate that is added to be converted to a detectable signal

what are the pros and cons of ELIZA test

pros: easy to preform and can test many samples quickly, inexpensive
cons: this test will give a number of false positives

what are the steps to the ELIZA test: detection of an antigen

1. primary antibody binds to wall

2. blocked agent is added

3. sample added; if antigen is present, it binds to antibody

4. unbound sample is washes away

5. antibody-enzyme conjugate is added

6. unbound secondary antibody-enzyme conjugate is washed away

7. substrate is added; if present, the enzyme converts substrate to colored products

what are the steps to the ELIZA test: detection of an antibody

1. antigen is attached to wall

2. a protein like gelatin is added to block the uncoated surface

3. patient antiserum is added; complementary antibody binds to antigen

4. enzyme-linked antibody is added to bound antibody

5. enzyme’s substrate is added, and reaction produces a visible color change

what is the western blot test

technique for detecting antigens in a complex mixture by separation of antigens by size (or detecting one antigen in a complex mixture of proteins)

describe the accuracy of the western blot test

can detect more types of antibodies and are less subject to misinterpretation than other tests (99.9% accurate)

what is nucleic acid tests (NAT)

researchers can detect genetic material by performing molecular techniques
- this can help understand if an individual is infected with a pathogen but does not show any signs or symptoms and is used to detect cancer genes

what is the southern blot vs northern blot

southern blot: detects specific DNA sequences from pathogen
northern blot: detects specific RNA sequences from a pathogen

what is the polymerase chain reaction (PCR)

technique used to create many copies of a DNA segment
- primers will bind to a specific region of the sample DNA genome and defines the region that will be amplified

what is RT-PCR

reverse transcription polymerase chain reaction (RT-PCR) is a variation of standard PCR that involves the amplification of specific RNA obtained from small samples

what is the cycle threshold of the RT-PCR

number of cycles in real time PCR assay that is needed for a positive reaction to be detected
- less cycles to detect a positive reaction = higher viral load
- higher cycles to detect a positive reaction = lower viral load