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Professor Leke Feb 2009
Professor Leke Feb 2009
INTRODUCTION
If parasites persist: Immune Response Modulated: If Re-infection enhanced, turned off, hypersensitivity.
INTRODUCTION
Tissue parasites Produce most pronounced IR. Ectoparasites, Intestinal worms less likely to evoke IR. IR: Dynamic event: can determine levels/types of Abs. Initial Ab Response: IgM later: IgG, ISgA, IgE. Helminths more likely than Protozoa to evoke IgE.
Complex antigenic composition of parasites cross reaction ags Lack of well standardized reagents and test procedures makes interpretation difficult. What does presence of Ab mean?
Abs present during active infection Abs stay after treatment Levels persist after infection has ceased false
positives
Direct Agglutination:
T- Cruzi epimastigotes. CATT = Card Agg - Test Tryps
2.
Principle: 1. Ab + Ag (low conc Ab) 2. Add Ag - coated RBC - agglutinated by uncombined free Ab
4. Complement fixation Add fixed Amt Ag to Test Serum (if ab+) Ag Ab complex Add C (known) Ag Ab C EA indicator cells if C available lysis. (RBC + sub agglutinating amt of Ab)
5. Immunodifussion in gels
Ouchterlony/Double diffusion = Ag & Ab moving PPt
Single Radial Immunodifusion / Mancini = Ig quantitative Immunoelectrophoresis = Ags separated using electrical charge PH Chosen = + proteins Neg Proteins move to + Cut Trough Fill with Ab-
6.
Electro-immunodiffussion (EID) : Electrically drive both ag and ab: counter current electrophoresis: Ab is + charged ; Ag is negative charged.
Rocket electrophoresis Laurell Used to quantitate Ag Also for Ab (single radial electro-Immunodiffusion: electrically driven) One dimensional pH chosen Ab imobile Ag, with negative charge Quant: Height of rocket proportional to ag conc.
7.
8.
Indirect: serum - labelled Ab can identify abs to different Ags in a single test
9. Enzyme Immunoassays
Enzyme linked immunoabsorbent assay = ELISA
Principle analogous to IFAT Couple enzyme to ab or ag Enzymes: Horseradish peroxidase., Alkaline phosphatase
Specific tests Schisto: Circumoval precipitin Test (COPT) of Olivier Gonsalesvery sensitive
1. ANTIGEN DETECTION.
ANTIGEN CAPTURE.
Use monoclonal / polyclonal Abs.
Involves the application of immunological techniques using antibodies (monoclonal) to detect malarial antigens, through antibody-antigen interactions. For malaria diagnosis, 2 important soluble antigens secreted by erythrocytic forms of the parasite in the blood are targeted:
Histidine rich Protein-2 (HRP-2),
Optimal test
-Antigen (pLDH binds antibody + gold particle -Ag-Ab- Gold particle complx migrate through test zone for Pf and Pv
5.
DNA from a selected region of a genome to be amplified a billion fold provided that part of its nucleotide sequence is already known. Known part of sequence used to design two synthetic DNA Oligonucleotides, one complementary to each strand of the DNA double helix and lying on opposite side of region to be amplified.