Rabbit antisera

Ramirez-Soto, D. and Poretz, R.D. (1991) The (l-3)-linked (X-L-fucosyl group of the IV-glycans of the Wistaria floribunda lectins is recognised by a rabbit antiserum. Carbohydate Research 213, 27-36. 

Antiserum Production The immunogen, carboxymethylmorphine-bovine-serum-albumin, is emulsified with equal volume of complete Freund s adjuvant. Initial immunization doses are injected into the New Zealand albino rabbits and later on this followed up with booster injections after a period of 6 weeks. The antiserum titer is determined with each booster dose injection and is duly harvested when the titre value is maximum. This is diluted suitably and employed in the radioimmunoassay. ... 

Immunization and Antibody Production The lypphilized immunogen obtained above is dissolved in normal saline and emulsified with equal volumes of complete Freund s adjuvant into a thick paste. Three New Zealand albino rabbits are immunized with the immunogen-paste through intradermal injections. The process is repeated twice at 2-weeks intervals followed by booster doses at monthly intervals. The antiserum is harvested when the plasma titer value is attained maximum. 

Preparation of Antiserum The barbiturate-bovine-serum-albumin conjugate is duly emulsified with an equal volume of complete Freund s adjuvant and New Zealand albino rabbits are subsequently im-... 

An emulsion of the hapten (i.e., conjugate) in normal saline is prepared by mixing with an equal volume of Freund s complete adjuvant. The prepared emulsion is injected subcutaneously into four different sites in New Zealand albino rabbits. Six weeks after the initial injection, all the animals are placed on a regimen of weekly booster shots. After a period of six months, antiserum from these animals are harvested and dilutions of 1 10,000 to 1 30,000 produced 50% binding or more and is employed in the RIA. 

The discovery of Lp(a) by Berg in 1962 (B6) relied on the production of rabbit antisera against beta-lipoprotein and on the selective absorption of these antisera with individual human sera. When certain human sera were used for absorption, the antisera retained precipitation capacity in radial immunodiffusion with 30-35% of individual human sera, which obviously contained a previous unknown antigen. The particle carrying the new antigen shared antigenic properties with beta-lipoprotein, but had an additional antigenic structure. This was evidenced from the only partial fusion of the precipitin bands formed between a positive human serum, the antibeta lipoprotein antiserum and the new absorbed antiserum. 

Figure 6.1. Fluorescence polarization immunoassay for theophylline. (A) Effect of theophylline rabbit antiserum ( ) and normal rabbit serum (A) on the fluorescence polarization of the theophylline-umbelliferone conjugate. (B) Fluorescence polarization of the theophylline-umbelliferone conjugate in the presence of varying concentrations of theophylline. (Reprinted from Ref. 1, with permission from Academic Press.)...

Preparation of Rabbit Serum and Antiserum. Four white New Zealand rabbits (2-3 kg) were first bled to obtain normal serum (NS), then injected subcutaneously with a total of 2.5 mg of either f-1, f-3, f-4, or f-5 in complete Freund s adjuvant. 

Figure 2. Double diffusion of cotton dust fractions and cotton bract extract against rabbit antiserum to dust (AD) and normal rabbit serum (NS). Conditions described in Material and Methods.

Figure 4. Double diffusion of extracts from cotton plant tissues and extract from flax against rabbit antiserum to cotton dust. Key. a AD,antisera to dust gin, gin trash If, leaf antigen stm, stem antigen cd, cotton dust antigen bet, bract antigen burr, burr antigen, b NS, normal rabbit serum, c fix, flax extract f-4, cotton dust antigen. Conditions described in Material and Methods.

Special thanks are given to Dr. Kolattukudy for providing the rabbit cutinase antiserum and purified cutinase used in this study. 

Dilute the respective antiserum, e.g., goat anti-(rabbit IgG) antiserum, to about 10 mg/ml with Soln. A and centrifuge or filter through a 0.45-pm membrane filter. 

Rinse a protein A column (4-5 ml bed volume 15 x 22.5 and 15 x 28.5 mm, respectively) with 40 - 50 ml of Soln. A, flow rate 1 ml/min. Dilute 2 - 5 ml of rabbit antiserum with the same volume of Soln. A and apply with the same flow rate to the column. After sample... 

Two recombinant allergens from olive pollen (Ole e 3 and Ole e 8) were produced in Arabidopsis and their relative biological activities assessed to be positive via calcium-binding assays. The immunological behavior of these recombinant allergens was equivalent to natural allergens, as demonstrated by their ability to bind to allergen-specific rabbit IgG antiserum and IgE from sensitized patients (Ledesma et al., 2006). 

An indirect competitive ELISA has been also developed for the determination of streptomycin and dihydrosticptomyciri in milk (24). Prior to the analysis, the milk sample was skimmed and treated with oxalic acid. The antiserum was raised in rabbits using streptomycin linked to a bacterial protein as the antigen. To perform the test, microtiter plates were coated with streptomycin, and antiserum and milk samples were mixed to be added in the wells where they were incubated for 1 h. Depending on the amount of residues in the sample, more or less antibody remained available for binding to the streptomycin coat. A pig antirabbit antibody-enzyme conjugate was subsequently added and incubated for 90 min. Using a suitable substrate, streptomycin and dihydrostreptomycin could be detected down to 1.6 ppb, whereas quantification could be made possible up to 100 ppb when samples were used undiluted. 

Mild conjugation reactions have been used in an enzyme-linked immunoassay for detecting cephalexin residues in milk, hen tissues, and eggs. The assay was a double-antibody separation procedure based on use of a rabbit antiserum... 

A competitive enzyme immunoassay for the quantification of ivermectin residues in bovine liver has also been reported recently (85). This method uses a polyclonal antiserum raised in rabbits against 5-O-succinoylivermectin-trans-ferrin conjugate. Cross-reactivity was demonstrated with doramectin, a member... 

At this stage, test the antiserum using an appropriate assay (see Note 5). If the antibody has the requirements for the use to which it will be put, up to three further bleeds on successive days may be performed. If the antiserum is unsatisfactory, i.e., the reaction is very weak, inject the rabbit again 1 mo after the test bleed, and again test-bleed 10 d after this injection. 

A rabbit may be bled every week provided that booster injections are administered every 8-12 wk. When this schedule is followed a large volume of antiserum can be collected from a single animal. 

A single rabbit can be expected to yield about 50 mL of antiserum, which should contain at least 10 mg of RIP-specific antibody... 

Fig. 1. Double label immunohistochemistry on rai liver. An acetone-fixed rat liver section was incubated with a polyclonal antiserum raised in rabbit to a hepa-tocyte cell surface protein (courtesy of Dr. S. Stamatoglou) and a mouse monoclonal antibody against a bile duct specific cytokeratin (courtesy of Dr. E. B. Lane). The hepatocyte protein was localized by use of a secondary peroxidase-conjugated antibody resulting in a red/brown product (thin arrow). The bile duct cytokeratin was identified by using an alkaline phosphatase-conjugated secondary antibody giving a blue color (thick arrow).

Add the reveal antiserum (for example, rabbit antigoat IgG alkaline phosphatase conjugate) at an appropriate dilution (1-1000 in Tris/saline), and incubate for 1 h at room temperature. 

The neutral FDPase and SDPase activities, which were present in the crude spinach extracts, were precipitated at lower ammonium sulfate concentration and could thus be separated from the specific alkaline FDPase. These activities appeared to be associated with the chloroplast fraction and did not require the presence of a divalent cation for activity. In crude extracts only the alkaline FDPase activity was inhibited by antiserum prepared by immunizing rabbits with the purified alkaline FDPase. The neutral FDPase was also active with ribulose diphosphate (RuDP) (98). 

Fig. 1.—Hapten Inhibition of the Precipitation of the Synthetic Antigen 8-Carboxy-octyl 4-Chloro-4-deoxy-0-D-galactopyranoside-BSA by Rabbit Antiserum Raised Against 8-Carboxyoctyl 0-D-Galactopyranoside-BSA (12 in Table I). [The haptens were the oj-earboxyalkyl /3-D-galactopyranosides having the aglycons (1) 8-carboxyoctyl, (2) 4-carboxybutyl, (3) 2-carboxyethyl, and (4) carboxymethyl (taken from Ref. 73).]...

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