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Freund's adjuvant

Freund's adjuvant is an immunological formulated as a water-in-oil to enhance the to co-administered s, primarily in preclinical research settings. It consists of two main variants: Complete Freund's Adjuvant (CFA), which includes , mannide monooleate as an emulsifier, and heat-killed (or M. butyricum), and Incomplete Freund's Adjuvant (IFA), which omits the mycobacterial components. Developed by Jules T. Freund in based on observations of heightened antibody production in mycobacteria-infected guinea pigs, it revolutionized experimental immunology by enabling potent induction of both humoral and cellular immunity at low doses. CFA is particularly noted for its ability to elicit strong Th1-biased responses, including delayed-type hypersensitivity and Th17 cell activation through mycobacterial stimulation of innate immunity and prolonged antigen depot effects at the injection site. In contrast, IFA promotes Th2-skewed humoral responses and is often used for booster immunizations following an initial CFA dose to minimize adverse effects. These adjuvants are widely applied in animal models for polyclonal antibody production, vaccine development studies, and induction of experimental autoimmune diseases, such as experimental autoimmune encephalomyelitis (a model for multiple sclerosis). Despite their efficacy, Freund's adjuvants are not approved for use due to significant concerns, including severe local , formation, tissue , and potential systemic from mycobacterial components in CFA. Early trials in the 1950s were halted after reports of adverse reactions, leading to their restriction to veterinary and applications under strict ethical guidelines. Ongoing explores modified versions or alternatives to harness their immunostimulatory benefits without the risks, underscoring their lasting influence on design.

Background

Definition and Composition

Freund's adjuvant is an immunopotentiator employed in immunological research to enhance immune responses to injected antigens through the formation of a stable water-in-oil . This encapsulates antigens in the aqueous phase within a non-metabolizable oil matrix, facilitating prolonged by creating a depot at the injection site that slowly releases the over time. The foundational composition of Freund's adjuvant revolves around a water-in-oil where the oil phase serves as the continuous medium. This phase primarily consists of , such as paraffin oil or light (e.g., Drakeol 6 or Bayol F), which is biologically inert and non-metabolizable, ensuring persistence in tissues. An emulsifier, mannide monooleate (known commercially as Arlacel A), is incorporated to stabilize the mixture, typically comprising 10-15% of the oil phase while makes up the remaining 85-90%. Antigens are suspended or dissolved in an , which is then emulsified with the oil phase in a standard 1:1 volume ratio to produce the final formulation. The resulting product exhibits a viscous, opaque, milky-white appearance due to the of aqueous droplets within the oil continuum, with droplet sizes generally in the micrometer range for optimal and slow release.

History and Development

Jules T. Freund (1890–1960), a Hungarian-born immunologist, pioneered the development of what became known as Freund's adjuvant during his tenure at the Department of , College of Medicine, in the late 1930s and early 1940s. Born in , Freund immigrated to the and established himself as a leading figure in applied , later serving as the first chief of the Division of Applied Immunology at the Public Health Research Institute of from 1943 to 1956. His work was motivated by observations of heightened immune responses in animals infected with , where the presence of mycobacteria appeared to amplify production against unrelated antigens. In 1937, Freund and colleagues published initial findings on the sensitizing effects of injecting tubercle bacilli suspended in paraffin oil, demonstrating enhanced formation and allergic responses in experimental animals. This laid the groundwork for research, but it was the 1942 collaboration with McDermott that formalized the adjuvant's core formulation: a water-in-oil incorporating heat-killed in , which dramatically boosted sensitization to horse serum in guinea pigs. This , later termed Complete Freund's Adjuvant (CFA), marked a significant advancement in techniques by providing prolonged release and potent stimulation of both humoral and cellular immunity. Subsequent studies in the and refined the , leading to the development of Incomplete Freund's Adjuvant (IFA) by omitting the mycobacteria to mitigate while retaining emulsifying . IFA was tested in human trials during the for diseases like and , showing promising enhancement, but its use was halted in the mid-1960s due to adverse reactions including local inflammation and formation. Freund's innovations, recognized with the 1959 Basic Medical Research Award for advancing procedures against infectious diseases, profoundly influenced modern design, inspiring safer emulsion-based systems still used in veterinary and experimental contexts today.

Formulation and Types

Complete Freund's Adjuvant

Complete Freund's Adjuvant (CFA) is a potent water-in-oil emulsion adjuvant formulated for maximal immunological stimulation, identical in base structure to the incomplete variant but augmented with heat-killed Mycobacterium tuberculosis (strain H37Ra) or M. butyricum at a concentration of 1–2 mg/ml. The emulsion comprises approximately 85% paraffin oil (or mineral oil), 15% emulsifier such as mannide monooleate (Arlacel A), and the mycobacterial suspension, creating a viscous mixture that traps antigens in depot form for prolonged release. Preparation of CFA for use involves sterile mixing of an equal volume (1:1 ratio) of the aqueous —typically sterile, , and pH-neutral—with the to avoid air bubbles and ensure stability. The mixture is then emulsified by vigorous agitation, commonly via the syringe method (repeatedly drawing the solution back and forth between two connected syringes fitted with a double-hub needle) or mechanical homogenization (e.g., using a bead-beater or at high speed for 1–2 minutes with cooling intervals on ). This process yields a stable, creamy water-in-oil , verified by its failure to disperse when a drop is placed in , with droplet sizes ideally under 1 μm for optimal ; commercial CFA vials are vortexed prior to mixing to resuspend the mycobacteria evenly. The distinctive immunostimulatory elements in CFA derive from the mycobacterial cell walls, notably muramyl dipeptide (MDP), a fragment that serves as a minimal active moiety mimicking bacterial signals to amplify innate and adaptive immunity. These components, absent in the base , enable CFA's superior potency over the incomplete form by driving robust Th1-biased responses, including enhanced production of IFN-γ and IgG2a antibodies, through recognition by receptors like NOD2. For storage, CFA is maintained at 2–8°C in a dark, sealed to preserve emulsion integrity and prevent microbial contamination, remaining stable for up to 12 months if unopened and handled aseptically; once mixed with , emulsions should be used immediately or refrigerated briefly, as separation can occur over time.

Incomplete Freund's Adjuvant

Incomplete Freund's (IFA) is a -in-oil formulated without mycobacteria, designed to prolong persistence through the formation of an oil depot at the injection site. This variant relies on the emulsified oil phase to slowly release the , thereby enhancing immune without the additional immunostimulatory components found in more potent formulations. The primary components of IFA include approximately 85% light , such as Drakeol 5NF or oil, and 15% mannide monooleate (Arlacel A) as the emulsifying agent, combined with an equal volume of the aqueous solution. Unlike formulations that incorporate heat-killed , IFA excludes such additives, resulting in a simpler that is heat-sterilized to ensure sterility with zero colony-forming units. Preparation involves vigorously mixing the oil-emulsifier base with the in sterile (pH 7.0–7.3) using a and needle until a stable, milky forms; stability is confirmed by testing a droplet's resistance to dispersion on the surface. Achieving fine droplet sizes, typically below 1 μm, is critical during emulsification to maintain integrity and prevent over time. IFA is commonly used for booster immunizations following an initial dose with a more inflammatory adjuvant, as it sustains antibody production while reducing local tissue reactions. This approach minimizes excessive inflammation during repeated administrations, making it suitable for protocols requiring multiple injections in animal models. Among its advantages, IFA presents a lower risk of granuloma formation compared to mycobacteria-containing variants, yet it remains effective in boosting humoral immunity by facilitating prolonged antigen exposure to B cells and T helper cells.

Mechanism of Action

Immunological Stimulation

Freund's adjuvant primarily functions by forming a subcutaneous depot at the injection site, which prolongs exposure through sustained release over several weeks. This water-in-oil traps the , preventing rapid clearance and allowing for gradual presentation to the . The depot effect enhances the of antigens, leading to more effective immune priming compared to soluble antigens alone. The stimulates innate immunity by recruiting antigen-presenting cells (APCs), such as dendritic cells and macrophages, to the injection site. These cells engulf and process the , facilitating its transport to draining nodes for broader immune activation. In the complete form, the inclusion of mycobacteria components further amplifies this recruitment and innate response. Freund's adjuvant enhances adaptive immune responses by increasing production, particularly IgG, and promoting T-cell . It influences the Th1/Th2 balance, with the complete variant favoring Th1 responses and the incomplete variant supporting Th2 profiles. Since its development in the 1930s, it has been widely used in experimental to enable detection of weak antigens and achieve polyclonal . This enhancement is evident in the magnitude and duration of titers and T-cell observed in preclinical studies.

Cellular and Molecular Pathways

Freund's complete adjuvant (CFA) primarily activates innate immune responses through components of heat-killed , which engage Toll-like receptors (TLRs) on antigen-presenting cells (APCs) such as macrophages and dendritic cells (DCs). Specifically, mycobacterial lipoarabinomannan and other components act as ligands for TLR2 and TLR4, initiating signaling cascades that culminate in the activation of the transcription factor via the adaptor protein MyD88. This pathway leads to the translocation of to the nucleus, promoting the expression of pro-inflammatory genes and enhancing APC maturation. The TLR-NF-κB activation in turn drives robust cytokine induction, creating a pro-inflammatory milieu essential for adaptive immunity. CFA stimulates the production of key cytokines including IL-1β, IL-6, TNF-α, and IL-12 from APCs, with IL-12 further promoting Th1 differentiation and TNF-α contributing to local inflammation and immune cell recruitment. Additionally, IL-23 production by activated DCs supports Th17 cell differentiation, facilitating IL-17-mediated responses that amplify autoimmunity in experimental models. These cytokines are secreted in a coordinated manner, with IL-6 and TGF-β synergizing with IL-23 to polarize naïve CD4+ T cells toward the Th17 phenotype. At the level of antigen processing and presentation, CFA enhances expression on s, improving the uptake and processing of co-administered antigens for presentation to + T cells. This is accompanied by upregulation of costimulatory molecules such as and on the DC surface, which provide the necessary signals (signal 2) for full T cell activation alongside TCR engagement (signal 1). Mycobacterial components, including heat-killed M. tuberculosis, directly induce this maturation phenotype in s, increasing , , , and levels through TLR-dependent pathways. A critical molecular component of CFA is muramyl dipeptide (MDP), derived from mycobacterial , which serves as a potent agonist for the intracellular . MDP binding to activates the receptor oligomerization and recruitment of RIPK2, leading to and MAPK pathway stimulation, which amplifies inflammatory signaling and production. Furthermore, -MDP engagement promotes activation, particularly involving , resulting in caspase-1 cleavage and processing of pro-IL-1β to its mature form, thereby intensifying the local inflammatory response. This -mediated amplification is essential for the adjuvant's potency, as -deficient models show diminished humoral and cellular responses to antigens emulsified in CFA. The oil-based in CFA contributes to a sustained molecular depot effect, where slow degradation of the vehicle prolongs the release of mycobacterial components and antigens at the injection site. This persistence facilitates ongoing production, such as (MCP-1), which recruits monocytes and DCs to the site, enhancing immune cell trafficking and chronic stimulation. The depot mechanism ensures that gradients, including IL-8 and MCP-3, are maintained over weeks, supporting prolonged APC activation and T cell priming without rapid clearance.

Applications and Effects

Use in Animal Research Models

Freund's adjuvant plays a central role in inducing experimental autoimmune encephalomyelitis (EAE), a widely used animal model for research. In this model, complete Freund's adjuvant (CFA) is emulsified with myelin antigens, such as (MOG) or myelin basic protein (MBP), and administered subcutaneously to like mice or rats, leading to T-cell-mediated inflammation and demyelination in the that mimics MS pathology. This approach has enabled detailed studies of autoimmune mechanisms and therapeutic interventions since the model's establishment in the mid-20th century. In arthritis research, Freund's adjuvant is essential for collagen-induced arthritis (CIA), a preclinical model of . CFA is mixed with and injected intradermally or subcutaneously into susceptible strains of mice (e.g., DBA/1) or rats (e.g., ), resulting in chronic joint inflammation, , and bone erosion that parallels human features. Additionally, adjuvant arthritis is induced solely by CFA injection without a specific , producing in rats to study innate immune responses and . Freund's adjuvant also features in models of type 1 diabetes, particularly in non-obese diabetic (NOD) mice, where CFA administration modulates disease progression by suppressing autoreactive T cells and preventing islet destruction, facilitating investigations into immune tolerance. Standard protocols typically involve an initial immunization with 100-200 μl of antigen emulsified in CFA, injected at multiple subcutaneous or intradermal sites, followed by booster doses using incomplete Freund's adjuvant (IFA) to sustain the response without excessive inflammation. Since its development in the , Freund's adjuvant has been indispensable for research, powering models like EAE and CIA that have advanced understanding of immune dysregulation. These models have notably contributed to the discovery and characterization of regulatory T cells, which suppress pathogenic responses in EAE, highlighting adjuvant-induced as a key tool for identifying immunotherapeutic targets.

Therapeutic Potential and Limitations

Freund's complete adjuvant (CFA) has demonstrated therapeutic potential in preclinical models of autoimmunity, particularly in type 1 diabetes, where it promotes beta cell regeneration through the induction of Th17 cells and interleukin-22 (IL-22) pathways. In non-obese diabetic (NOD) mice, adjuvant immunotherapy with CFA upregulates regenerating (Reg) genes, such as Reg2, in pancreatic islets, leading to reduced insulitis, increased insulin secretion, and partial restoration of normoglycemia. This effect is mediated by IL-22-producing Th17 cells, which enhance beta cell proliferation and survival while preventing apoptosis, independent of MyD88 and IL-6 signaling pathways. These findings highlight CFA's ability to modulate autoimmune destruction toward regenerative outcomes in rodent models of diabetes. Beyond , CFA enhances efficacy against weak antigens in infectious disease and cancer models. In studies, CFA boosts Th1 and Th17 immune responses, increasing interferon-gamma (IFN-γ) production and splenocyte proliferation when combined with immunogens like p27, thereby promoting activation and protective cellular immunity. Similarly, in tumor models, CFA's water-in-oil facilitates prolonged , upregulating co-stimulatory molecules on antigen-presenting cells and eliciting robust cytotoxic T responses to epitopes with inherently low . These enhancements have been observed in animal systems, underscoring CFA's role in amplifying humoral and cellular immunity for challenging antigens. Despite these benefits, CFA's therapeutic applications are severely limited by its propensity to induce unwanted and excessive , hindering translation to human use. The adjuvant's mycobacterial components can trigger off-target immune reactivity, such as responses to non-relevant antigens, potentially exacerbating autoimmune conditions rather than resolving them. In models like experimental autoimmune encephalomyelitis (EAE), CFA confounds results by independently activating glial cells and skewing toward Th1 dominance, which misrepresents disease pathogenesis and impairs pain assessments critical for clinical relevance. Its inflammatory reactions, including granulomatous lesions and severe local pain, render it unsuitable for human trials, as evidenced by restricted use to preclinical settings. Post-2020 developments have focused on synthetic alternatives that mimic CFA's immunostimulatory effects without its toxicity, alongside continued exploration in veterinary vaccines. Synthetic adjuvants like glucopyranosyl (GLA) derivatives and cyclic dinucleotides (CDNs) activate (TLR4) and cGAS-STING pathways, respectively, to drive Th1-biased responses for and cancer vaccines with improved safety profiles. However, ethical concerns arise from over-reliance on CFA in immune tolerance models, as it induces unnecessary animal suffering through inflammatory pain and skews interpretations of tolerance mechanisms, potentially invalidating translational insights into human .

Regulation and Safety

Regulatory Guidelines

The use of Freund's adjuvant in humans has been prohibited by the U.S. (FDA) since the mid-1960s due to its association with severe adverse reactions, including sterile abscesses observed in clinical trials. It is also not suitable for human use according to guidelines from the (EMA), established in 1995. No containing Freund's adjuvant have been approved for human use by the FDA or EMA. In animal research, the use of Freund's adjuvant, particularly complete Freund's adjuvant (CFA), is strictly regulated and requires oversight by Institutional Animal Care and Use Committees (IACUCs) in the United States, as well as accreditation bodies like the Association for Assessment and Accreditation of Laboratory Animal Care International (AAALAC). Administration is limited to subcutaneous or intraperitoneal routes, with maximum volumes of 0.1 ml per subcutaneous site or 0.2 ml intraperitoneally for , to minimize and distress. CFA is typically restricted to the initial dose, with incomplete Freund's adjuvant (IFA) preferred for boosters, and all protocols must include scientific justification for its necessity over milder alternatives. Under international standards, the Directive 2010/63/EU classifies procedures involving CFA as requiring detailed justification due to their potential for moderate to severe effects on animals, emphasizing the 3Rs principle (, reduction, refinement) and encouraging the use of synthetic or less invasive alternatives where possible. This directive mandates that ethical review bodies assess the harm-benefit balance, with CFA's application limited to cases where no suitable substitutes exist. Historically, trials of Freund's adjuvant in the 1950s, including for vaccines, were halted in the mid-1960s following reports of adverse events such as sterile abscesses and prolonged induration at injection sites. All research protocols involving Freund's adjuvant must include mandatory documentation of adverse events, such as or ulceration, with prompt reporting to attending veterinarians and IACUCs to ensure ongoing monitoring and protocol adjustments.

Adverse Effects and Risks

Freund's adjuvant, particularly Complete Freund's Adjuvant (CFA), induces significant local reactions at the injection site, including acute and chronic , formation, ulceration, abscesses, and tissue sloughing due to the persistence of the oil emulsion, which can last for months. These granulomatous responses often involve alterations and can lead to persistent nodules. In studies with rabbits and mice, subcutaneous or intramuscular injections of CFA resulted in severe pathological changes at the site, with behavioral indicators of discomfort more pronounced after secondary doses. Systemic effects from Freund's adjuvant include widespread granuloma dissemination to organs such as the lungs, , liver, and kidneys, triggered by migration of the oil components and mycobacterial elements. In domestic , administration led to hypercalcemia and compromised renal function in multiple cases, with elevated serum attributed to activation. Oxidative stress markers, such as increased thiobarbituric acid reactive substances () in liver and muscle tissues, along with elevated plasma tumor necrosis factor-alpha (TNF-α), have been observed in mice, indicating broader inflammatory damage. The adjuvant's potent immunostimulatory properties can inadvertently induce autoimmune-like responses, including chronic inflammation that mimics conditions such as . In BALB/c mice, CFA exposure resulted in as a prominent adverse outcome, accompanied by disrupted profiles and heightened . Such unintended autoimmune induction arises from the adjuvant's ability to provoke excessive and prolonged immune activation beyond the targeted response. Animal welfare concerns are substantial, as Freund's adjuvant causes notable and distress, particularly with footpad injections that impair and naturalistic behaviors like burrowing and wheel running. In experimental models, these reactions necessitate rigorous pain scoring, monitoring for severe distress, and predefined criteria to prevent unnecessary suffering. Behavioral alterations in mice, including reduced activity post-injection, underscore the welfare impact. Long-term risks encompass chronic potentially leading to carcinogenicity, with rare instances of persistent nodules and even injection-site sarcomas reported in . The oil's prolonged retention contributes to ongoing tissue irritation and , raising concerns for sustained health impairments in research animals.

References

  1. [1]
    The ABC of clinical and experimental adjuvants—A brief overview
    Freund's adjuvant is an example for water-in-oil emulsions. This adjuvant is based on the observation of Jules Freund, that guinea pigs infected with ...Fig. 1 · 5.3. Mf59 (type B Adjuvant) · 6.3. Toxin-Derived Adjuvants...
  2. [2]
    Guidelines on Adjuvant Use in Rats and Mice - Research A-Z
    Freund's Complete Adjuvant (FCA): FCA is a water-in-oil emulsion of mineral oil, mannide monooleate (a surfactant), and heat-killed or avirulent Mycobacterium ...
  3. [3]
    Old and new adjuvants - PMC - NIH
    Emulsion based adjuvants. In 1930, Jules Freund developed the first deliberately added adjuvant that enhanced cellular immunity, a water-in-mineral oil ...
  4. [4]
    Complete Freund's adjuvant induces experimental autoimmune ...
    Complete Freund's Adjuvant (CFA) emulsified with an antigen is a widely used method to induce autoimmune disease in animal models, yet the contribution of ...
  5. [5]
    https://animalcare.research.northeastern.edu/wp-content/uploads/2022/05/Polyclonal-Antibody-ProduCiton-Final-2022.docx
  6. [6]
    An Overview of Vaccine Adjuvants: Current Evidence and Future ...
    Complete Freund's adjuvant (CFA) includes in its structure heat-killed mycobacteria, which enhances the stimulation of immune responses and is currently used ...
  7. [7]
    Peptide emulsions in incomplete Freund's adjuvant create effective ...
    Sep 9, 2022 · Water-in-oil emulsion incomplete Freund's adjuvant (IFA) has been used as an adjuvant in preventive and therapeutic vaccines since its ...
  8. [8]
    Adjuvants and Antibody Production: Dispelling the Myths Associated ...
    Jul 1, 2005 · Paraffin oil, a crude mineral oil, and the emulsifying agent mannide monooleate (Arlacel A®) were used in the original formulation of Freund's ...
  9. [9]
    https://www.sigmaaldrich.com/US/en/product/mm/344291
  10. [10]
    Standardization of Antigen-Emulsion Preparations for the Induction ...
    ... emulsions, Complete Freund's Adjuvant, emulsification. Introduction. Mouse ... The ratio of aqueous solution/adjuvant was always kept 1:1 (v/v). If other ...
  11. [11]
    Sensitization to Horse Serum by Means of Adjuvants - Sage Journals
    First published April 1942. Request permissions. Sensitization to Horse Serum ... Freund's adjuvant is a classic of vaccine adjuvants and the basis of ...
  12. [12]
    Jules Freund - The American Association of Immunologists
    Freund's scientific contributions were many, but he is most remembered for developing a technique for enhancing immune responses with the adjuvant that bears ...
  13. [13]
    Freund's Adjuvant - an overview | ScienceDirect Topics
    Freund's Adjuvants. Since the original descriptions of its formulation and efficacy (Freund, 1951, 1956; Freund and McDermott, 1942; Freund et al., ...
  14. [14]
    (PDF) Freund's Adjuvants - ResearchGate
    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 ...Missing: original | Show results with:original
  15. [15]
    Freund's adjuvant - Lasker Foundation
    Feb 27, 2021 · Freund's studies to preventive medicine may be discerned in several important developments. For example, in early investigations of poliovirus ...Missing: history | Show results with:history
  16. [16]
    https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/335/648/f5881pis.pdf
  17. [17]
    CFA | Complete Freund's Adjuvant for research - InvivoGen
    Complete Freund's Adjuvant (CFA) consists of heat-killed Mycobacterium tuberculosis in non-metabolizable oils (paraffin oil and mannide monooleate).
  18. [18]
    [PDF] Guidelines for the Use of Adjuvants in Research - NIH OACU
    Guidelines for the Preparation of Complete Freund's Adjuvant: 1. The mycobacteria in CFA is re-suspended by vortexing or shaking the ampule or vial. The CFA ...
  19. [19]
    Muramyl dipeptide and its derivatives: peptide adjuvant in ... - NIH
    In 1974, MDP was discovered to be the minimal structure required for the efficacy of Freund's Complete Adjuvant (FCA), one of the most potent and widely used ...
  20. [20]
    Muramyl Dipeptide - an overview | ScienceDirect Topics
    It is the smallest structural component of the cell wall that still retains adjuvant activity and is one of the active components in Freund's complete adjuvant.
  21. [21]
    Freund's Adjuvant - an overview | ScienceDirect Topics
    This is an oil-based agent with droplets of aqueous peptide contained within a surrounding oil phase. This facilitates a depot effect at the vaccine site, ...Nature Of Antigens · 4.10 Adjuvants · 6 Adjuvants In Experimental...
  22. [22]
    [PDF] No-Waste™ Freund's Complete Adjuvant - G-Biosciences
    Upon arrival, store at 4°C. Freund's Complete. Adjuvant is stable for up to 1 year if stored correctly and handled with aseptic techniques. Do not freeze ...
  23. [23]
    [PDF] CFA - Complete Freund's Adjuvant - NET
    CFAVax consists of a mixture of mineral oil and emulsifier in a ratio of 85% v/v oil and 15% v/v emulsifier. Each 10. mL CFA vial contains 10 mg mycobacterium.<|control11|><|separator|>
  24. [24]
    https://www.mpbio.com/us/freunds-incomplete-adjuvant
  25. [25]
    [PDF] Florida State University Antibody Production - Adjuvants
    The primary purpose of an adjuvant is to enhance the immune response to a particular foreign protein (antigen) of interest. In the context of antibody ...
  26. [26]
    https://www.scbt.com/p/freundprimes-incomplete-adjuvant
  27. [27]
    Vaccine adjuvants: mechanisms and platforms - Nature
    Jul 19, 2023 · 2). Subsequently, in the 1940s, Freund and his colleagues developed water-in-oil emulsions, which led to the creation of Freund's adjuvants. ...
  28. [28]
    Modes of action of Freund's adjuvants in experimental models of ...
    Dec 1, 2001 · Freund had suggested three categories of action mechanisms: 1) prolong[ation of] the presence of antigens at the site of injection, 2) more ...
  29. [29]
    An Overview of Vaccine Adjuvants: Current Evidence and Future ...
    Complete Freund's adjuvant (CFA) includes in its structure heat-killed mycobacteria, which enhances the stimulation of immune responses and is currently used in ...
  30. [30]
    Role of Toll-like Receptors in Adjuvant-Augmented Immune Therapies
    Freund's complete adjuvant (FCA) consisting of dead mycobacteria ... RIP1 is an essential mediator of Toll-like receptor 3-induced NF-kappa B activation.
  31. [31]
    Th17 cytokines and Vaccine Induced Immunity - PMC
    Most autoimmune models deliver antigens in adjuvants such as Complete Freund's Adjuvant (CFA) [52]or pertussis toxin [39] to induce the inflammatory disease ...
  32. [32]
    Critical role of M. tuberculosis for dendritic cell maturation to induce ...
    The expression of interferon-gamma (IFN-gamma), Interleukin-12p40 (IL-12p40), costimulatory molecules CD40, CD80 and CD86 and MHC class II were up-regulated in ...
  33. [33]
    Modulation of adaptive immunity by different adjuvant-antigen ... - NIH
    Sep 27, 2008 · Muramyl dipeptide (MDP) is a component of the peptidoglycan polymer and was shown to be an active but low-toxicity component of complete ...
  34. [34]
    The ever-expanding function of NOD2: autophagy, viral recognition ...
    ... MDP is the 'minimum' effective component of complete Freund's adjuvant that promotes adaptive immune responses [45–47]. It has been demonstrated that MDP ...
  35. [35]
    Vaccine Adjuvants | Review - InvivoGen
    MDP triggers the activation of NOD2 and the NLRP3 inflammasome. ... Incomplete Freund's adjuvant | Th2 response. 1 product from $76. VIEW · Vaccination ...Mechanisms Of Adjuvants · Alum & Emulsions · Prr Ligands
  36. [36]
    Complete Freund's adjuvant as a confounding factor in multiple ...
    Feb 15, 2024 · Complete Freund's adjuvant (CFA) is used as a standard adjuvant for the induction of experimental autoimmune encephalomyelitis (EAE).
  37. [37]
    Experimental Autoimmune Encephalomyelitis as a Testing ...
    Sep 16, 2010 · EAE can be induced by inoculation with central nervous system (CNS) proteins or peptides emulsified in complete Freund's adjuvant.
  38. [38]
    Applicability and implementation of the collagen-induced arthritis ...
    The collagen-induced arthritis (CIA) mouse model is the first choice and the most widely used because it may be generated rapidly and inexpensively.
  39. [39]
    Native type II collagen-induced arthritis in the rat. I. Incidence ... - NIH
    An acute inflammatory arthritis has been induced in 76% of rats injected intradermally with native bovine type II collagen emulsified in Freund's complete (CFA) ...
  40. [40]
    Adjuvant Arthritis - an overview | ScienceDirect Topics
    Adjuvant arthritis (AA) is defined as a model of arthritis induced in certain rat strains by the administration of complete Freund's adjuvant (CFA) or other ...
  41. [41]
    Prevention of type I diabetes in NOD mice by adjuvant immunotherapy
    We report that a single injection of complete Freund's adjuvant (CFA) given at an early age (5 wk) prevented the appearance of diabetes and greatly increased ...
  42. [42]
    Effects of different doses of complete Freund's adjuvant on ...
    Dec 8, 2021 · This study aimed to compare the effects of different high doses of CFA in rat to closely mimic nociceptive and inflammatory parameters of rheumatoid arthritis ...
  43. [43]
    Live Imaging of Immune Responses in Experimental Models of ...
    EAE models have contributed to the understanding of immunoregulatory processes during the pathogenesis of MS, and CD4+CD25+FoxP3+ regulatory T (Treg) cells ...
  44. [44]
    Revisiting Freund's incomplete adjuvant for vaccines in the ...
    Aug 7, 2025 · Due to emerging adverse effects such as local skin reactions, abscesses, inflammation or granulomas at the injection site, IFA is not allowed ...
  45. [45]
    Common Ingredients in FDA-Approved Vaccines
    Jan 12, 2024 · When evaluating a vaccine for safety and effectiveness, FDA considers adjuvants as a component of the vaccine; they are not approved separately.Missing: Freund's | Show results with:Freund's
  46. [46]
    [PDF] Guideline on adjuvants in vaccines - European Medicines Agency
    Mar 25, 2004 · Similarly, complete Freund's adjuvant, although being one of the most powerful adjuvants known, is not suitable for human use. Interest in ...Missing: ban | Show results with:ban
  47. [47]
    Standard on Use of Adjuvants for Antibody Production - UNC Policies
    All adjuvants used in animal research must be approved by the IACUC, and use of adjuvants that could induce a severe reaction must be scientifically justified.
  48. [48]
    Boston University Institutional Animal Care and Use Committee ...
    Jun 3, 2025 · The use of adjuvants must be described and scientifically justified in the IACUC protocol, to include volume, site and frequency of administration.
  49. [49]
    Use of Complete Freund's Adjuvant (CFA) and Other ... - UW Sites
    Discontinuance of the use of CFA has been recommended, and use of alternative adjuvants is strongly encouraged (see resources to identify alternatives below).
  50. [50]
    [PDF] IACUC Policy: Complete Freund's Adjuvant (CFA) Use
    The use of adjuvants in animal research requires careful consideration, particularly Complete Freund's. Adjuvant (CFA). This compound can cause severe side ...Missing: AAALAC | Show results with:AAALAC
  51. [51]
    [PDF] IACUC Policy on the Use of Freund's Adjuvant in Laboratory Animals
    If Freund's complete adjuvant must be used, it should be limited to the initial immunizing dose. Use of two or more doses of the complete adjuvant is rarely ...
  52. [52]
    Applying refinement to the use of mice and rats in rheumatoid ...
    Replacing or avoiding animal use should be a principal goal, as required in many legislations including European Directive 2010/63/EU (European Commission 2010) ...
  53. [53]
    [PDF] Severity assessment Framework - Caring for animals - CroLASA
    The Commission established an Expert Working Group (EWG) for the assessment of severity of procedures to facilitate the implementation of Directive 2010/63/EU ...
  54. [54]
    [PDF] The Critical Review of Methodologies - European Commission
    New diagnostic methods continue to identify hitherto unrecognized sensitizing fragrance chemicals making the total fragrance burden for EU citizens unchanged.
  55. [55]
    Reporting of Adverse Events - University of Washington
    Aug 21, 2025 · Regardless of cause, all adverse events must be promptly reported to the AV. The AV will work with the reporting individual and with the Office ...
  56. [56]
    Announcements | IACUC | Research Integrity & Compliance | USF
    The PI of such an approval is required to report to the IACUC the number of animals used in research, and any adverse events occurring during the research.<|control11|><|separator|>
  57. [57]
    Assessment of side effects induced by injection of different adjuvant ...
    We evaluated the side effects induced by injection of Freund's adjuvant (FA) and alternative adjuvants combined with different antigens.
  58. [58]
    Immunologic responses and adverse reactions to Freund's ...
    Seven cats developed extensive granulomatous reactions at injection sites, lymph nodes, and multiple visceral organs including lungs and brain. Persistent ...
  59. [59]
    Freund´s complete adjuvant effect on BALB/c mice - PubMed
    Despite high efficacy, it is not used in human vaccination due to number of adverse effects. Arthritis can be an example of a typical adverse consequence.
  60. [60]
    Behavioral outcomes of complete Freund's adjuvant-induced ... - NIH
    The greatest negative effects can be observed in naturalistic behaviors such as burrowing and wheel running. Subgroup analyses most often revealed animal ...Missing: adverse | Show results with:adverse