Freund adjuvant

Nitro substituted carboxylic acids, N,N-dicyclohexylcarbodiimide (DCC) and N-hydroxysuccinimide were obtained from the Aldrich Chemical Co. (Milwaukee, WI). Avidin-labeled horseradish peroxidase, biotinylated anti-rabbit IgG, ovalbumin ( M.W. 45,000), bovine serum albumin (M.W. 66,000) Freund s complete adjuvant, Freund s incomplete adjuvant, and o-phenylenediamine were purchased from Sigma Chemical Co.(St. Louis, MO). All solvents were reagent grade. DME (1,2-dimethoxyethane) was dried by distillation from sodium-potassium amalgam/benzophenone ketyl. 

Water-in-oil emulsions such as Freund s complete adjuvant, Freund s incomplete adjuvant. The complete adjuvant contains QdX- aDlQ6. Mycobacterium tuberculosis bacteria whereas the incomplete does not. Both have the basis of an immune modulator, e.g., branched glucose polymers or methylated bovine serum albumin (Pierce s AdjuPrimeTM). 

Polyclonal Antibodies against FORL r purified polygalacturonase were raised in white rabbits. For the first immunization 200 jxg of purified protein in 300 jxl of distilled water was mixed with 200 1 of PBS and 500 n of complete Freund s adjuvcmt and injected intramusculary into the leg. Two subsequent intramuscular injections, each containing 300 fig of protein in 1 ml of incomplete Freund s adjuvant were given at 1 month intervals. Finally, the rabbit was bled 1 week later. The antisera, separated from blood by incubation at 37 "C, were stored in 1 ml fractions at -20 C. 

Compared to wild-type mice, in TRPVl gene-deleted (—/—) mice, complete Freund s adjuvant evokes significantly less oedema, hyperalgesia and arthritis score [149]. 

Two goats were immunized three times during the first 2 weeks with 1 mg of the antigen emulsified in 1 ml of Freund s complete adjuvant at several subcutaneous sites near regional lymph centers. Booster injections of 3 mg of antigen were administered at monthly intervals. The animals were bled 7 days after each boost. After several months of immunization, the titer and affinity of the antibody response was judged sufficient for use. 

AIA, adjuvant-induced arthritis CFA, complete Freund s adjuvant CIA, collagen-induced arthritis DTH, delayed-type hypersensitivity LPS, lipopolysaccharide. 

Materials. Microspherical PGG glucan (Adjuvax, Alpha-Beta Technology, Worcester, MA) was prepared from Saccharomyces cereviseae strain R4 cells (11). Zymosan, cytochrome c (cyt c), bovine serum albumin (BSA), yeast alcohol dehydrogenase (ADH), Complete Freunds Adjuvant (CFA) and Incomplete Freunds Adjuvant (IFA) were purchased from Sigma Chemical Co. (St. Louis, MO). 

These results show that the covalently crosslinked Adjuvax formulations were superior to the physically entrapped Adjuvax formulations. In addition, the Adjuvax crosslinked formulations were as effective in stimulating antibody titers as Freund s Adjuvant. Furthermore, animals immunized with Adjuvax did not experience local inflammatory reactions or granuloma formation which was observed with all CFA/IFA immunized animals. 

Figure 4. Comparison of Freund s Adjuvant to Adjuvax formulations in stimulating antibody response to P55 oligopeptide antigen. Relative antibody titers between adjuvant groups at day 27 were determined by measuring the absorbance at 450 nm of a 1 500 dilution of anti-P55 immune sera by ELISA.

Jongwon C, Keun H, Suk-Hwan K, Kyung-Tae L, Hyeong-Kyu L. Hee-Juhn P. Kalopanaxsaponin A from Kalopanax pictus, a potent antioxidant in the rheumatoidal rat treated with Freund s complete adjuvant reagent. J Ethnopharmacol 2002 79 113-118. 

Fig. 15 Induction of cellular immunity by subcutaneous immunization with OVA-encapsulating y-PGA-Phe nanoparticles. Mice were subcutaneously immunized one time with OVA alone (10 pg), 10 pg of OVA and 100 pg of NPs (OVA-NPs), 10 pg of OVA and 100 pL of complete Freund s adjuvant (OVA + CFA), or PBS (control). Splenocytes were obtained from the immunized mice on day 10 after the immunization and stimulated with the OVA peptide. The number of IFN-y-producing cells was measured by enzyme-linked immunospot assay. SFU spot forming units...

A corollary to the use of cBSA as a carrier protein is that its increased immune response often abrogates the use of complete Freund s adjuvant, which is a source of concern because of its potential side-effects in animals. A relatively innocuous mixture with alum is usually all that is required as adjuvant to result in good antibody production. 

The adjuvanticity of oil emulsions was first recognized in the early 1900s. However, the first such product to gain widespread attention was Freund s complete adjuvant (FCA), developed in 1937. This product essentially contained a mixture of paraffin (i.e. mineral) oil with dead mycobacteria, formulated to form a water-in-oil emulsion. Arlacel A (mannide mono-oleate) is usually added as an emulsifier. 

Freund s incomplete adjuvant (FIA) is a similar product. It differs from FCA in that it lacks the mycobacterial component and, consequently, displays somewhat lesser adjuvanticity. The mode of action of FIA is largely attributed to depot formation. The mycobacterial components in FCA have additional direct immunostimulatory activities. Although it is one of the most potent adjuvant substances known, FCA is too toxic for human use. 

AP immunotherapy has been used in both prevention and treatment trials in mutant mice. Both Ap immunization (with Freund s adjuvant) and passive transfer of Ap antibodies reduce levels of AP and plaque burden in... 

In contrast to certain oil emulsions (including Incomplete Freund s Adjuvant), the liposphere approach uses pharmaceutically acceptable biodegradable constituents. 

FIGURE 7.7 (See color insert) Adoptively transferred D011.10 transgenicT cells can be identified by expression of CD4+ and KJ-126 in spleen cell suspension from Balb/c mice after ovalbumin (OVA) immunization. Balb/c mice were injected iv with D011.10 spleen cells containing 3-5 x 1 06CD4+KJ-126+ cells and immunized by intraperitoneal injection of 2 mg OVA emulsified in complete Freund s adjuvant 2 days later. OVA immunization increases the frequency of KJ+T cells and alters the expression of various surface molecules consistent with T cell (Tc) activation. 

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