Immunization methods

Gershon P.D., Khilko S., Stable chelating linkage for reversible immobilization of oligohistidine tagged proteins in the BIAcore surface plasmon resonance detector, J Immun Methods 1995 183 65-76. 

Fig. 1. Schematic drawing of various immune methods for localization of antigens. (A) Direct method. (B) Indirect method. (C) Immunogold with silver enhancement. (D) Peroxidase-antiperoxidase method. (E) Avidin-biotin system.

This is an immunization method in which nitrocellulose-bound protein is employed and in which neither DMSO nor Freund s adjuvant are used, in contrast to the method described by Knudsen (4) It is equally applicable for soluble and membrane proteins. 

Second, methods for the characterization of complex antisera are difficult. Antisera to E. coli protein mixtures have been developed with impressive spectra of reactivity using conventional immunization methods (6,22-23). An exact assessment of the spectrum of antibody reactivity is often limited, however, by the resolution of the analytical methods used. Counter immunoelectrophoresis is limited by the relatively low sensitivity of detection and resolution for complex mixtures of reacting species. One dimensional silver stained SDS-PAGE and immunoblotting provides sensitive detection limits but lacks resolution. Therefore, methods which have a high degree of resolution and sensitivity are required to best compare potential improvements in the production of antibodies to minor components in the mixture. 

FIGURE 3.1.2 The principle of antigen determination by the immune methods with avidin-biotin signal amplification, (a) Labeled avidin-biotin assay (LAB assay) (b) Bridged avidin-biotin assay (BRAB assay) and (c) Avidin-biotin complex method (ABC assay). (From jQdrychowski, L., Pol. J. FoodNutr. Sci., 12/53(SI 2), 32, 2003b. With permission.)... 

Analytical methods include immune methods (ELISA) and liquid chromatography/mass spectrometry (LC/MS). Gastric contents can be assayed and ricin can be detected in blood and body fluids by radioimmunoassay and LC/MS (Darby et al, 2001 Mouser et al, 2007). 

Parenteral Route. Parenteral vaccination remains the immunization method of choice for most antigens because it provides more effective immune response than do any other routes of vaccination in most cases. Every years millions of people receive inactivated influenza vaccine by parenteral administration. Subcutaneous vaccination with inactivated influenza vaccine is known to induce simultaneous immune responses in the blood and upper respiratory tract of subjects. The immune response, i.e., the increase in the number of influenza virus-specific antibody-secreting cells in peripheral blood and tonsils, increased rapidly to reach a peak within 1 week after vaccination.Parenteral vaccination of a DNA vaccine encoding glycoprotein D of herpes simplex virus type 2 resulted in systemic cellular and humoral responses. The mucosal humoral responses generated by intramuscular and intradermal vaccination were comparable with those obtained by mucosal vaccination. The DNA vaccine was able to... 

Folkersen J, Tiesner B, Westergaard JG, Grudzinskas JG (1985) J Immun Methods 77 45... 

Botchkareva AE, Eremin SA, Montoya A, Manclus JJ, Mickova B, Pavel R, Fini F, Girotti S. Development of chemiluminescent ELISAs to DDT and its metabolites in food and enviroiunental samples, J Immun Methods 2003 283 45-57. 

Levraud, J. P., Colucci-Guyon, E., Redd, M. J., LutfaUa, G., and Herbomel, P. (2008) In vivo analysis of zebrafish innate immunity. Methods Mol. Biol. 415, 337-363. 

Blomberg K. (1994) Simultaineous Measurement of Natural Killer cell cytotoxicity against each of three different target cell lines. J.Immun. Methods. 168 267-273. 

Bohlen H, Manzke O, Engert A, Hertel M, Hippler-Altenburg R, Diehl V, and Tesch H. (1994) Differentiation of cytotoxicity using target cells labelled with europium and samarium by electroporation. J. Immun. Methods. 173 55-62. 

Laukkanen M-L, Orellana A, Keinanen K. (1995) Use of genetically engineered lipid-tagged antibody to generate functional europium chelate-loaded liposomes. J. Immun. Methods. 185 95-102... 

Dressendorfer R,A, Jorg-Martin H, Gerzer R, and Strasburger C, (1995). A Non-isotopic immunoassay for guanosine 3 5 -cyclic monophosphate using a cyclic GMP-biotin conjugate as tracer. J.Immun.Methods. 

This chapter describes the production of anti-serums, the purification of antibodies, and the most important immunological screening technologies. For monoclonal antibodies, I refer you to Methods Enzymol. I (1986) 121, Hybridoma Technology and Monoclonal Antibodies. I also touch only briefly upon the newer in-vitro immunization methods in the following (with friendly consultation from Dr. Rose-John, Mainz). 

E. Zacco, M. I. Pividori, X Llopis, M. del Valle, and S. Alegret, Renewable Protein A modified graphite-epoxy composite for electrochemical immunosensing]. Immun. Methods, 286,35-46 (2004). 

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