Adjuvant Potency
The success of any vaccine adjuvant formulation is dependent upon fulfillment of several requirements. The most important of these are potency, tolerable reactogenicity, and pharmaceutical feasibility. In order to be protective, the antigen adjuvant formulation must have a specific potency for generation of the appropriate immune function. It is useful to classify adjuvants in terms of their performance regarding the correlation of immunity historically established for a variety of disease...
David C Neujahr and David S Pisetsky 1 Introduction
DNA is a complex macromolecule with immunological properties that depend on base sequence. Although mammalian DNA is immunologically inert, DNA from bacteria has potent immunostimulatory properties that result from short sequence motifs called CpG motifs or immunostimulatory sequences (ISS). These motifs, which have the general structure of two 5' purines, an unmethylated CpG motif, and two 3' pyrimidines, occur much more commonly in bacterial DNA than mammalian DNA because of two main factors...
Limulus Lysate Assay
Commercial Limulus polyphemus amoebocyte lysate (LAL) test kits, e.g., either Sigma E-Toxate, multiple test vial system sensitive to 0.005-0.5 endotoxin units (EU) mL Cat. 210-2, or M.A. Bioproducts' LAL test system with a reagent which will detect 0.25 ng mL of FDA reference endotoxin with the addition of 1.0 ng mL of Escherichia coli O 111 B4. Sigma, Cat. 50-505U. 2. Pyrogen-free water for making dilutions of the standard, e.g., Endotoxin-free water Sigma Cat. 210-7. 3. All glassware must be...
Subunit Vaccines
It is becoming increasingly apparent that the natural immune response generated by infection with an organism often does not represent the optimal protective response against that organism (12). Subunit vaccines, consisting of specific components of the pathogen, can be used to direct the immune response to targets that are protective (13). Strain variation of immunologically important sites can be covered by the use of mixtures of peptides, proteins, or nucleic acids that code for them. In...
Hemolysis Test
A New Zealand white (NZW) rabbit or equivalent. 2. Heparinized bleeding set, e.g., Vacutainer (Becton and Dickinson, Rutherford, NJ). 3. Sodium chloride (0.85 (w v) Sigma, St. Louis, MO). 5. Cyanmethemoglobin standard, Merck Ltd., Darmstadt-Mannheim, Germany, Cyanmethemoglobin standard for photometric determinations of hemoglobin, 1.0 mL in 200 mL distilled water. Cat. 36210P or Hemoglobin standard, Sigma, Cat. 525-A. 6. Hematocrit and bench centrifuge. 8. Drabkin's reagent (Merck) this reagent...
Granulocyte Macrophage Colony Stimulating Factor GMCSF
GM-CSF was originally identified as a survival and growth factor for hematopoietic progenitor cells and a differentiation and activating factor for granulocyte and monocyte cells (12). Recently, it has been shown to enhance the maturation of dendritic cell precursors (13). It is a glycoprotein of approximately 16,000 Dalton mol wt (127 amino acids aa in the human) containing two disulfide bonds (14). It is secreted by activated T cells, macrophages, fibroblasts, and endothelial cells. The...
Jrg Kreuter 1 Introduction
Nanoparticles are solid particles ranging in size from 1 to 1000 nm (1 m). They consist of macromolecular materials and can be used therapeutically or prophylactically, for example, as adjuvants in vaccines or drug carriers, in which the active principle (drug or biologically active material) is dissolved, entrapped, or encapsulated, or to which the active principle is adsorbed or chemically attached (1-3). One of the first areas of application of nanoparticles was their employment as adjuvants...
General Elisa Protocol
Dilute coating monoclonal antibodies to a final concentration of 1 g mL in PBS pH 8.5. 2. Add 100 L of diluted coating antibody to each well of a 96-well Immulon 2 ELISA plate. Seal the plate with parafilm to prevent evaporation. Allow antibody to coat for 12-18 h at 4 C (see Note 6). 3. Wash plate four times with PBS 0.1 Tween-20. An automated plate washer is useful in this and subsequent wash steps. Slap the plate against a pile of paper towels to remove residual fluid. 4. Block the wells by...
Release Assays for MF59C1
MF59C.1 is a well-defined emulsion produced to preestablished release specifications. The emulsion bulk and final single-dose adjuvant are analyzed using a battery of assays in accordance with Chiron's standard operating procedures. Key assays include visual appearance, pH, mean particle size, and number of large particles per milliliter for quality, squalene, polysorbate 80, and sorbitan trioleate concentrations by high-performance liquid chromatography (HPLC) procedures for content and,...
Advantages of Adjuvants
Vaccine adjuvants influence the immune response to our benefit in one or more ways (see Table 3). The ability of adjuvants to influence so many parameters of the immune response greatly complicates the process of finding an Modulators of Vaccine Adjuvant Effects Intraspecies Genetic Variation effective adjuvant. This is because our knowledge of how any one adjuvant operates on a cellular level is insufficient to support a completely rational approach for matching the vaccine antigen with the...
Methods 31 Formulation
The SAF emulsion formulation was typically manufactured as a twofold concentrate. Prior to use, the emulsion was diluted with a twofold concentrate Components of the 2X SAF Emulsion Concentrate Potassium phosphate monobasic, NF 0.0184 Sodium phosphate dibasic, anhydrous, USP 0.11 Water for injection, USP q.s. 100.0 of threonyl-MDP and antigen. The threonyl-MDP was formulated in an isotonic buffered solution at a pH that was optimal for its stability. The emulsion formulation, as manufactured,...
Immune Responses to TCI
CT and other related ADP-ribosylating exotoxins such as heat-labile entero-toxin from E. coli (LT) are in themselves strong immunogens when given by routes other than TCI. In our early experiments, we noted that high levels of anti-CT antibodies could be rapidly induced after one application of CT to the Fig. 1. Kinetics of the serum IgG antibody response to CT (A), DT (B), or BSA (C) in animals immunized (0 wk) and boosted (4 and 8 wk) by the transcutaneous route with CT (100 g), DT (100 g)...
SAF Components and Chemistry
Threonyl-muramyl dipeptide (threonyl-MDP), Syntex (Palo Alto, CA). Threonyl-MDP is comprised of a muramyl sugar derivative and two nonaromatic amino acids linked by amide bonds. Its solubility is greater than 600 mg mL in aqueous solution over a wide pH range. Its solubility in nonpolar solvents, such as chloroform, methylene chloride, and hexane, is less than 3 g mL. The apparent octanol water and methylene chloride water partition coefficients were found to be 0.0044 and 0.00017,...
References
H., Lindblad, E. B., Bizzini, B., Ben-Efraim, S., and Gupta, C. K. (1993) Adjuvants a balance between toxicity and adjuvanticity. Vaccine 11, 293-306. 2. Gupta, R. K. and Siber, G. R. (1995) Adjuvants for human vaccines current status, problems and future prospects. Vaccine 13, 1263-1276. 3. Gupta, R. K., Rost, B. E., Relyveld, E., and Siber, G. R. (1995) Adjuvant properties of aluminum and calcium compounds, in Vaccine Design The Subunit and Adjuvant Approach...
By Covalent Coupling of the Antigen see Notes 3 4 and
Antigens (hepatitis A, hepatitis B, diphtheria toxoid, tetanus toxoid, and sPf66 malaria synthetic peptide) are bound to IRIV surface by phosphatidylethanola-mine (PE) whose free amino group allows a covalent coupling. 2. PE is dissolved in methanol and 0.1 (v v) triethylamine is added. The solution is then mixed with y-maleimidobutyric acid N-hydroxy-succinimide ester (GMBS) (Pierce Chemical Company, Rockford, IL) (ratio PE GMBS 2 1) which is previously dissolved in dimethylsulfoxide (DMSO)....
Determination of Surface Properties of Microparticles Using Scanning Electron Microscopy SEM
A highly porous or irregular surface of the microparticles can lead to a high burst release or dumping of the antigen. Therefore, it is important to evaluate the surface properties of the microparticles under a scanning electron microscope (see Note 10). 1. 10 mg of dried microparticles are suspended in 500 L of distilled water to yield a viscous suspension. 2. 100 L of this suspension is added on surface of metal stubs used for SEM. 3. The stubs are air-dried and then sputter coated with...
Conclusions
PMMA nanoparticles are polymeric particulate adjuvants for vaccines. These nanoparticles can easily be manufactured in a reproducible manner in the described particle sizes and with specific surface properties. Scale-up of the production process is also facile (20). PMMA is a material with a good safety record that has been used in humans for more than 50 yr. PMMA nanoparticle adjuvants achieved good antibody responses and good protection against challenge with a number of antigens....
Discussion
Manufacturers are expected to provide safety data sheets for their products and these will confirm the lack of toxicity of the product (see Note 4), but the addition of the vaccine candidate antigen may also alter the overall reactivity of the complete vaccine. Therefore, it is unwise to rely completely on manufacturer's specifications alone. Before evaluating the possible toxicity of a new adjuvant formulation, it is assumed that the investigator will have checked for either the innate...
Notes
Table 1 summarizes factors affecting adsorption of antigens onto aluminum adjuvants. Adsorption of antigens onto aluminum adjuvants depends heavily on electrostatic forces between adjuvant and antigen (3,6,7,19). Other interactions including hydrophobic, van der Waals, and hydrogen bonding contribute to the adsorption of antigens on aluminum adjuvants. However, these forces may not suffice to cause adsorption of antigen if the same charge or electrostatic repulsive force is present on antigen...
Induction of Allergy to Nonvaccine or Food Proteins
This is a particularly important test when examining the suitability of an adjuvant for inclusion in an oral vaccine, as there could be a reaction to food proteins (23). With the interest in the oral route as a means of stimulating mucosal immunity, there is a possibility that an adjuvant could induce an allergic response to dietary proteins. In this study, both lactalbumin and gluten failed to elicit an IgE response in the presence of the original Freund's Complete or Incomplete Adjuvants (FCA...
Preparation of ISCOMs Hydrophobic Interactions
Amphipathic Antigens (Native or Lipidated) in Detergent 1. Make a preparation of the antigen in detergent. 2. Mix antigen, lipids, and saponin according to Table 4 and adjust the volume with PBS. Incubate for 1-2 h at room temperature prior to extensive dialysis against PBS. 3. Dialyze against 3-5 changes of buffer (24-48 h), the first 24 h at 20-25 C then at +4 C. 3.2.2. Antigen in 3-8 M urea (Gua-HCl, DMSO, EG, and so on) 1. If the antigen is solubilized in, e.g., urea, mix antigen,...
Production of a Commercialized Influenza Virosome Vaccine see Note
At the Swiss Serum and Vaccine Institute, Berne, influenza seed virus solution is inoculated into 11-d-old embryonated hens' eggs from flocks under veterinary control. The inoculated eggs are incubated for a further 50-60 h at 33-35 C, depending on the strain. During this period, they are illuminated for a second time (after 40 h) to eliminate dead eggs. After incubation, the eggs are cooled overnight at 1-4 C they are then opened under laminar flow and the allantoic fluid is aspirated into...
Selection of Vaccine Adjuvant Candidates for Clinical Trial
The decision to begin human trials of vaccines and adjuvanted vaccines is complex and depends on a number of criteria (85). 1. The vaccine adjuvant candidate must address a public health need, and it must be a logical means to prevent or treat the disease of interest. 2. The vaccine adjuvant must have been designed with a sound scientific rationale. 3. There must be an expectation of safety, as discussed in the section above on safety. 4. There must be animal studies demonstrating the...
DC Dialysis Method
This method for encochleation involves the removal of detergent from a solution of lipid and material to be encochleated by dialysis against a buffer containing calcium. This method has been applied to the formulation of vaccines containing membrane proteins (although protein recovery is lower than the LC method), and DNA plasmids. 1. Material to be encochleated is added to (or purified in) a solution containing a detergent in a high salt buffer (e.g., 2 Octyl P D glucopyranoside in extraction...
Prospects for the Future
Because it is not clear which (if any) animal species or strain correlates with human immunity (54), products that show promise in an animal system need to be tested in humans for both safety and immunogenicity. Virosomes have been extensively given to humans of all ages in the context of vaccine formulations. Therefore, such efforts should prove exciting for the successful application of the molecular approach to new and improved vaccines. There are several ways to increase adjuvant activity...
Summary and Conclusion
Interest in vaccine adjuvants is intense and growing, because many of the new subunit vaccine candidates lack sufficient immunogenicity to be clinically useful. In this chapter, I have emphasized modern vaccine adjuvants injected parenterally or administered orally or intranasally with licensed or experimental human vaccines in volunteers. The terms adjuvant, carrier, vehicle, and adjuvant formulation are defined. Every adjuvant has a complex and often a multifactorial immunological mechanism,...
Studies in Humans
Two large clinical trials have compared adjuvanted HIV vaccines (62,71) and adjuvanted malaria vaccines (28) in healthy young adult volunteers. These trials illustrate results that can be obtained from comparative adjuvanted vaccine trials in volunteers using similar clinical protocols. In a phase 1, doubleblind, randomized, placebo-controled trial in healthy adults, 50 g of HIV gp120 was combined with one of seven adjuvants (62). The summary of side effects caused by these vaccines and...
Assurance of Clinical Safety
The most critical concern in the development of postalum adjuvants has been the demonstration of safety for the large populations who receive the Summary of Subjects Receiving MF59 and Vaccine Antigens through March 1999 in Chiron-Sponsored Programs Summary of Subjects Receiving MF59 and Vaccine Antigens through March 1999 in Chiron-Sponsored Programs vaccine. Adjuvants, by definition, increase immunological responsiveness, which may result in immune reactivity to epitopes other than those...
Jason A Neidleman Gary Ott and Derek OHagan 1 Introduction
Heat-labile enterotoxin (LT) from Escherichia coli and Cholera toxin (CT) from Vibro cholerae are known to be potent mucosal immunogens. These toxins have 80 sequence homology and a similar tertiary structure (1-4), and both elicit potent serum IgG and mucosal IgA responses (5,6). Moreover, both also serve as excellent adjuvants for coadministered antigens. However, they are toxic in their native state and both produce accumulation of intestinal fluid and watery diarrhea (7). LT is the cause of...
Preparation of ISCOMs Antigen Lipidation Techniques
Hydrophilic antigens that need introduction of an hydrophobic moiety in order to incorporate into ISCOMs can be readily lipidated (attachment of a lipid tail) at e-amino groups (in principle lysine side chains) using, e.g., hydroxysuccinimide esters of fatty acids or at carboxy groups (aspartic and glutamic acid residues) using carbodiimide and aminogroup containing lipids. Alternatively thiol-binding lipids can be attached to naturally occurring thiols (cystein) or at chemically introduced...
Plasmids and Yeast Strain
Plasmid pOGS40 contains a C-terminal truncated p1 gene (amino acids 1-381) in a multicopy yeast plasmid (Fig. 1). A BamHI site immediately 5' to the stop codon allows additional coding sequence to be inserted. Plasmids pOGS 40, 41, and 42 have the BamHI site in reading frames 1, 2, and 3, respectively, but are otherwise identical. These three plasmids contain the yeast LEU2 marker that will transform a leu2 auxotroph to leucine independence. The p1 gene is under the control of the strong,...
Derek T OHagan and Manmohan Singh 1 Introduction
Over the last 20 years, the adjuvant effect achieved through the association of antigens with polymeric microparticles has been repeatedly demonstrated (1,2). Encapsulation of antigens into microparticles, including submicron particles (nanoparticles), promotes their entry into lymph nodes and provides a high local concentration of antigen over an extended time-period. Micro-particles also promote the interaction of encapsulated antigens with antigen presenting cells (APCs), e.g., macrophages....
Local Reactions
The most frequent adverse side effect associated with adjuvanted vaccines is the formation of local inflammation with signs of swelling and erythema, and symptoms of tenderness to touch and pain on movement. Such reactions occur more frequently in preimmune individuals, or after repeated immunization (24). The inflammation is thought to be the result of formation of inflammatory immune complexes at the inoculation site by combination of the vaccine antigen with preexisting antibodies and...
Reaction Profile of Freunds Adjuvants
Freund's adjuvants are used in priming immunizations. In boosters, FIA can be used with antigen or antigen alone can be administered in saline. Interestingly, it was shown by Paraf and coworkers in mouse studies, that injection of human serum albumin in saline prior to a stimulating injection of HSA in FCA, could completely suppress the antibody reponse (35). In rabbits, injection of HSA with Freund's adjuvant was able to elicit a significant antibody response in the newborn individual, which...
Comparative Vaccine Adjuvant Trials 111 Animal Studies
Modern studies have compared up to 24 investigational adjuvants individually mixed with one antigen in a single protocol reviewed by Edelman (88) . The single protocol controls for confounding test variables, such as antigen, dose, schedule, animal species, and immunological assays. These variables make comparisons between two or more separately conducted studies difficult, if not impossible. When adjuvants provide equally good immunogenicity in such comparison trials, adjuvant choice may...
Mechanisms of Adjuvant Action
To date, most subunit vaccines are poor antigens, whether or not they are natural products, recombinant products, or synthetic peptides. Subunit antigens fail for a variety of reasons, such as incorrect processing by the immune system, rapid clearance, stimulation of inappropriate immune response, and lack of critical B-cell or T-cell epitopes. Potentially, some of these failures can be overcome by administering subunit antigens with adjuvants. It should be remembered, however, that the best...
Reinhard Glck 1 Introduction
Immunization is the most effective defense mechanism against microbial infections today. Although highly effective vaccines are currently available for a number of infectious diseases, vaccine formulations can still be improved in a number of important areas. Issues of safety, stability, delivery, and combining vaccines to several pathogens need to be addressed. For many diseases, a greater understanding of microbial pathogenesis and the basis for protective immunity is still needed. The...
Robert Edelman 1 Introduction
Adjuvants have been used to augment the immune response to antigens for more than 70 years. Ramon first demonstrated that it was possible to increase levels of diphtheria or tetanus antitoxin by the addition of bread crumbs, agar, tapioca, starch oil, lecithin, or saponin to the vaccines (1). In this chapter, an overview is provided of modern vaccine adjuvants as background for more detailed discussions of promising adjuvants in chapters to follow. After a more general discussion of adjuvants...
Qs21 Mass Spectrometry
Pharmaceutical Properties and Storage QS-21 is manufactured by Aquila Biopharmaceuticals, Inc., and is supplied as a lyophilized, odorless, white powder, which is stored at < -20 C until formulation (up to 3 yr postmanufacture). It is > 98 pure by RP-HPLC, has < 10 endotoxin units mg QS-21, < 10 colony-forming units (CFU) mg QS-21, and < 5 residual moisture. 2.1.2. Spectroscopic Identification Positive ion fast-atom bombardment mass spectrometry can be employed for...
ISCOM Technology
The classical procedure for ISCOM formation is to mix antigens and saponin with detergent-solubilized cholesterol and phospholipid (8). ISCOMs are formed when detergent is removed by dialysis, ultrafiltration, or ultracentrifu-gation (9-11). However, not all antigens spontaneously incorporate into ISCOMs using standard procedures even if they are hydrophobic also, many From Methods in Molecular Medicine, Vol. 42 Vaccine Adjuvants Preparation Methods and Research Protocols Edited by D. T....
Preclinical and Phase I Clinical Trial Design Issues 101 US Food and Drug Administration Regulations
No detailed or specific guidelines exist in the United States for assessing the safety of adjuvant preparations for use in humans. Only two guidelines refer to adjuvants. The first guideline formally issued by the FDA, which includes adjuvanted vaccines (86), refer to tests of the final container lot of all biological products. These FDA standards are paraphrased in Table 7 for ease of understanding. It is unclear if adjuvants, such as QS-21, which are added to the vaccine immediately before...
Materials
Protein antigen The handling of the antigen before incorporation is important. The antigens most suited for ISCOM formulation are hydrophobic or amphipathic. Such antigens must be purified in the presence of a detergent to prevent micelle or aggregate formation because pure protein micelles or aggregates are very difficult to dissociate to make their hydrophobic part accessible for hydrophobic interactions. There are many examples where antigens that theoretically should incorporate well, in...
Duncan E S Stewart Tull 1 Introduction
There are no officially recognized regulations for the design and toxicity testing of adjuvants or adjuvant formulations the former are also referred to as immunomodulators and immunopotentiators. At the Immunological Adjuvants and Vaccines meeting held in Greece in 1988, however, immuno-adjuvant researchers discussed experimental toxicological tests that might be used to monitor new immunomodulators (1). The usefulness of these tests for the range of immunomodulators and adjuvant formulations...
The Use of MF59 as a Vaccine Adjuvant
The MF59 emulsion adjuvant was developed with the objective of generating a broad spectrum of recombinant vaccines for human use. The specific aim was to elicit neutralizing titers in humans significantly greater than those obtainable with the alum adjuvants in common usage. An extensive body of preclinical efficacy data has been obtained based on ELISA titers obtained with a variety of subunit antigens in a spectrum of animal model systems. A summary of this data is shown in Table 2. Data are...
Measurement of the Toxicity of Adjuvants
Cytotoxicity Assay in Cultured Monolayers of Human or Animal Cell Lines This type of assay has the great merit of reducing the number of animals which must be used to comply with standardized toxicity tests. The MTT assay (13) was adapted for determining cell survival and proliferation by a number of workers (14-16) with different cell types and toxins.The assay compares favorably with other similar systems (17), is less time-consuming and objective than microscopic examination of cells,...
Immunopotentiating Reconstituted Influenza Virosomes
Twenty years after the discovery of the immunological adjuvant properties of liposomes (25) and the ensuing multitude of related animal immunization studies (26), liposomes as adjuvants have come of age (27,28) with the first liposome-based vaccine against hepatitis A being licensed for use in humans. Vaccines based on novasomes (nonphospholipid biodegradable, pausilamellar vesicles formed from single-chain amphiphiles, with or without other lipids) have also been licensed for the immunization...
Composition of Mineral Oil Adjuvants
As mentioned, the classical Freund's incomplete adjuvant consists of a mixture of mineral oil and emulsifier in a ratio of 85 v v oil and 15 v v emulsi-fier. This mixture will then be mixed with an equal volume of aqueous solution of antigen and subsequently emulsified prior to use. Ratios of 90 oil and 10 emulsifier have also been described. Both components will be discussed below. The oil component of mineral oil adjuvants is normally light mineral oil of a highly purified quality. Some...
Protein Cochleates
Cochleates as carriers for protein and peptide antigens effectively induce antibody- and cell-mediated immune responses. Protection from lethal and Summary of Cochleate Vaccine Studies Summary of Cochleate Vaccine Studies Intestinal intraepithelial lymphocytes infectious dose challenge with viruses administered parenterally and mucosally has also been achieved (see Table 1 for summary) (14-19, and manuscripts in preparation). For example, cochleate vaccines containing the glycoproteins and...
Rajesh K Gupta and Bradford E Rost 1 Introduction
Aluminum compounds, including aluminum phosphate (AlPO4), aluminum hydroxide (Al(OH)3), and alum precipitated vaccines, historically referred to as protein aluminate, are currently the most commonly used adjuvants with human and veterinary vaccines (1-6). These adjuvants are often referred to as alum in the literature, which is misleading because (1) two most widely used adjuvants from this group, aluminum hydroxide and aluminum phosphate, have very different physical characteristics (7) and...
Sarah C Gilbert 1 Introduction
Adjuvants are available to promote the generation of antibodies to an antigen following immunization. However, many of these adjuvants do not enhance priming of cytotoxic T lymphocytes (CTL). The reason for this lies in the existence of two alternative antigen processing pathways, leading to stimulation of CD4+ T cells, and, in turn, to the generation of antibodies, or stimulation of CD8+ CTL. In general, exogenous proteins enter the antigen presenting cell (APC) by endocytosis. Peptides...
Final Vaccine Formulation
A variety of antigens including HSV-2 gB gD, HBV, HIV gp120 and or p24, CMV gB, and influenza hemagglutinin (HA) have been formulated with Fig. 4. Long-term particle size stability of the three representative MF59C.1 bulk lots at 2-8 C. Fig. 4. Long-term particle size stability of the three representative MF59C.1 bulk lots at 2-8 C. the adjuvant emulsion MF59C.1 in one of two formats either in single containers or in separate vials (admixed prior to administration and therefore referred to as...
Gregory M Glenn Tanya Scharton Kersten and Russell Vassell 1 Introduction
Transcutaneous immunization (TCI), the introduction of antigens using a topical application to intact skin, is a new technology that has both practical and immunological merits. Practically speaking, a needle-free method of vaccine delivery will decrease the risk of needle-borne diseases, reduce the complications related to physical skin penetration, improve access to vaccination by eliminating the need for trained personnel and sterile equipment, and provide a simple means for multivalent or...
Historical Background
The first evidence of the use of oil emulsions as adjuvants in vaccine formulations dates back to 1916. At that time, Le Moignic and Pinoy immunized mice with heat-inactivated Salmonella typhimurium in an emulsion of water and vaselin oil, using lanolin as an emulsifier (1). The oil emulsions as adjuvants, however, did not receive much attention before Jules Freund and coworkers (2,3) decades later combined a paraffin (mineral) oil emulsion and heat-killed mycobacteria to produce an extremely...
Examples of Modern Vaccine Adjuvants Used in Animals and
Agents listed in Table 1 are examples of the many varieties of immuno-potentiators used during the past 30 years. The majority of adjuvants are being developed and tested by industry. The list of adjuvants is incomplete, because I have not conducted an exhaustive literature search, because the results have appeared in abstracts in nonindexed publications, and because many studies are proprietary. The adjuvants marked by an asterisk in Table 1 have completed trial in man, or they are now...
Erik B Lindblad 1 Introduction
Freund's adjuvants are water-in-mineral oil emulsions (W O emulsions) without heat-killed mycobacteria added (Freund's incomplete adjuvant) or with heat-killed mycobacteria added (Freund's complete adjuvant). Freund's adjuvants are, in a way, historic adjuvants and for many researchers the first and most obvious association to the word adjuvant at all. The adjuvants have been used extensively in experimental immunology owing to their high efficacy, and Freund's incomplete adjuvant has been used...
Preclinical Evaluation of Vaccines Adsorbed onto Aluminum Compounds
To achieve consistency in manufacture of aluminum adjuvants, it is important to characterize these adjuvants for physicochemical and adjuvant properties. Though aluminum adjuvants have been used widely with human vaccines, there are still a number of unanswered questions about their mechanism of action (see Note 5), such as optimal size of the gel particles, degree of adsorp tion of antigens onto adjuvant, amount of gel used as a dose for humans and animals (see Note 3) and even type of...
Terry Ulrich 1 Introduction
Interest in new methods of potentiating the immune response against vaccine antigens has increased considerably over the past decade. In part, this interest is in response to vaccine initiatives that have established aggressive goals for improving existing vaccines and for developing much-needed new vaccines. Many of the candidate vaccine antigens being developed as part of this effort are synthetic or recombinant subunit structures that are often poorly immunogenic and as such, are unable to...
General Adjuvant Properties
QS-21 stimulates strong antibody- and cell-mediated responses in animals. QS-21 has been shown to stimulate strong antibody responses to T-dependent protein antigens in mice 3-5 , guinea pigs 6 , rhesus monkeys 7 , and baboons 8 . Antibody responses induced by QS-21-adjuvanted vaccines were shown to be higher than antibody responses with aluminum hydroxide-adjuvanted vaccines and were similar to those obtained with Freund's com-plete-adjuvanted FCA vaccines 3,6 . QS-21 is also a strong adjuvant...
General Structure and Physicochemical Properties
QS-21 was shown by 1H and 13C-NMR and fast-atom bombardment-mass spectrometry to be an acylated 3,28-o-bisdesmodic triterpene saponin of molecular weight 1990 16 . The structure of QS-21 is shown in Fig. 4. QS-21 is anionic at neutral pH, because of the carboxyl group on glucuronic acid. It is insoluble in aqueous solution in the protonated form, but soluble in the sodium salt form where QS-21 forms micelles in solution. The critical micellar concentration of QS-21 was estimated to be 26 M in...
Aluminum Phosphate
Like aluminum hydroxide, aluminum phosphate is the most widely used adjuvant with routine human vaccines. Most of the vaccine manufacturers throughout the world prepare this adjuvant in-house. Usually, antigens are adsorbed onto a preformed gel, which can be made by several methods two methods are described in this chapter . Adsorption of antigens is also carried out on freshly prepared aluminum phosphate gel 23 . In situ adsorption of antigens onto aluminum phosphate is preferable to aluminum...
Center for Biologics Evaluation and Research Cber Fda
The CBER, FDA, Rockville, MD is responsible for regulating vaccines and other biologics in the United States. In addition to meeting the general standards before public release Table 7 , each vaccine and adjuvant are tested for safety on a case by case basis, preferably with the help and guidance of the CBER as noted before. Such guidance, informal in nature but quite helpful, was published in 1993 in response to the needs of HIV-1 vaccine development 52 . The principles laid down by that...
Vaccine Formulation
Mix the sterile stock solution of QS-21 with a sterile stock of soluble antigen with the dose of each adjusted to the optimum for the animal to be immunized. A typical effective QS-21 dose is 10-20 g for mice, 25-50 g for guinea pigs or rats, and 50-100 g for rabbits, rhesus monkeys, and baboons. The volume of the vaccine is then adjusted to the final formulation volume 0.2 mL for mice, 0.5-1.0 mL for larger animals with sterile saline or sterile-buffered saline. Although QS-21 is a...
