Study Antigen targets Methodology Conventional diagnostic methods Diagnostic accuracy metrics Clinical utility Summary of main results Improvements in diagnostic accuracy or clinical decision-making
Hasler et al. [8] Not explicitly stated Immunofluorescent staining techniques used to detect autoantibodies Clinical appearance, histopathology, ultrastructural analysis Not explicitly provided, but IF helped confirm diagnosis in cases where clinical presentation was equivocal Immunofluorescence provided a valuable diagnostic tool to differentiate between PV, bullous pemphigoid, and other similar bullous disorders Immunofluorescence helped confirm the diagnosis of PV or bullous pemphigoid in 4 patients with oral lesions, where conventional methods were equivocal Immunofluorescence allowed differentiation between PV, bullous pemphigoid, and other similar bullous disorders that can present with oral lesions
Donatsky et al. [9] Unkeratinized buccal human oral mucosa from RAS patient Immunofluorescence staining, microscopic examination, absorption experiments to ensure specificity - Compared antibody endpoint titres between controls and RAS patients, but did not report quantitative accuracy metrics Supports autoimmune aetiology for RAS RAS patients had higher antibody titres to adult human oral mucosa compared to controls.
No difference in titres using RAS vs control oral mucosa as antigen.
Absorption experiments confirmed specificity of positive IF 		Provides evidence for autoimmune mechanism in RAS, but does not directly improve diagnostic accuracy or clinical decision-making
Torabinejade et al. [10] Immune complexes 1. Periapical lesions were frozen, sectioned, and examined using
ACIF technique.
2. Complement and anti-complement
were used at 1 : 20 dilution.
3. Positive and negative controls were used 		Histopathological examination of formalin-fixed sections - ACIF technique was able to detect immune complexes in 23 out of 25 periapical lesions, suggesting a role of immune complexes in the pathogenesis of periapical lesions 23 out of 25 periapical lesions were positive for antigen-antibody complexes using ACIF technique.
No staining was observed in the 2 lesions diagnosed as periapical scar tissue 		ACIF technique was more sensitive than the DIF technique used in previous studies, allowing for better detection of immune complexes in periapical lesions. This could provide insights into the pathogenesis of these lesions
Acosta et al. [11] Intercellular substance of stratified squamous epithelium Epithelial cells collected from oral mucosa by scraping.
Cells spread directly on slides or washed and cytocentrifuged onto slides.
DIF staining performed 		Histology. IIF for serum intercellular antibodies - DIF on oral cytological smears may be useful for diagnosing PV IgG deposition was observed on cytological smears from oral lesions in all patients with active PV.
No fluorescence was seen in smears from pemphigus patients in remission, patients with other oral diseases, or healthy controls.
Washing the cells prior to IF improved specificity by reducing non-specific fluorescence 		DIF on oral cytological smears may provide a more specific and less invasive diagnostic method for PV compared to traditional histology and IIF
Daniels et al. [12] Immunoglobulins, complement components, and other protein substances within affected tissues Cryostat sectioning, incubation with diluted antisera, washing, mounting, and fluorescence microscopy examination History, clinical features, histopathology, and clinical follow-up Not reported (qualitative assessment of DIF patterns in relation to diagnosis) DIF findings can establish diagnosis of pemphigus, pemphigoid, lichen planus, and LE; absence of characteristic patterns can help rule out these conditions DIF can provide distinguishing patterns to aid in the diagnosis of chronic ulcerative oral mucosal diseases DIF findings can strengthen or rule out diagnoses, thereby improving diagnostic accuracy and clinical decision-making for chronic ulcerative oral mucosal diseases
Fine et al. [13] BMZ antigens Includes tissue processing and potentially antigen retrieval techniques, but specifics are not provided Clinical examination and histopathological analysis Not reported Enhanced understanding of the disease mechanism, potential impact on diagnosis Immunofluorescence and immunoelectron microscopy provide detailed insights into the pathology of cicatricial pemphigoid Better visualization of antigen-antibody complexes and complement deposits in tissue samples, aiding in the differentiation of CP from other similar conditions.
Firth et al. [14] Fibrinogen, C3, Clq, IgA, IgG, IgM One portion of the biopsy was processed for routine histopathology (H&E staining), while another portion was frozen, sectioned, and stained with the fluorescent-labelled antibodies for DIF examination. Clinical features, routine histopathology (H&E staining) Concordance between clinical, histopathological, and DIF diagnoses DIF was considered an "integral step" in the diagnosis of OMLP, as it contributed to the diagnosis in 13 cases and helped exclude OMLP in 9 cases where the clinical and/or histopathological findings were equivocal. In 116 (70%) cases, all three diagnostic modalities (clinical, histopathological, and DIF) agreed on the diagnosis of OMLP.
In 13 (7.9%) cases, the diagnosis could only be established by DIF. In 9 (5.6%) cases, DIF helped exclude a diagnosis of OMLP when the clinical and/or histopathological findings were equivocal.
DIF was non-contributory in 27 (16.3%) cases. 		The study concludes that DIF was an "integral step" in the diagnosis of OMLP, as it provided additional diagnostic information not available from clinical and histopathological evaluation alone.
Lodi et al. [15] Epithelial antigens on monkey oesophagus substrate IIF technique using pre-fixed sections of monkey oesophagus Clinical and histological features of OLP Not reported Determine the nature and frequency of circulating antibodies to epithelial antigens in the sera of HCV-positive patients with OLP There was a significant association between the concomitance of OLP and HCV infection and the presence of circulating antibodies to epithelial antigens Some patients with HCV-associated OLP may have circulating antibodies to epithelial antigens, although their precise etiological role in the development of this disease in HCV infection remains unknown
Yih et al. [16] Immunoglobulins and complement component within affected gingival tissues Biopsy of gingival lesions, with one half processed for H&E histology and the other for DIF using frozen sections Clinical presentation and routine histological examination DIF analysis, in combination with histology, was able to provide a definitive diagnosis in 65 out of 72 (90.3%) DG cases Correct diagnosis is critical for determining appropriate treatment and follow-up, as DG can be caused by a variety of immunological and idiopathic conditions with different prognoses Improvements in diagnostic accuracy or clinical decision-making DIF analysis, combined with histology, provided a more accurate diagnosis compared to clinical presentation and routine histology alone, which frequently cannot differentiate between the various DG-causing conditions
Bhol et al. [17] α6 integrin-BMZ Sera of patients and determination of their effects on normal human buccal mucosa in organ culture Clinical symptoms (erosions, vesicles, or bullae in the oral cavity), histological findings (subepithelial cleft with mixed cell infiltrate) Not specified Identification of target antigens and understanding their effects on buccal mucosa Identify the target antigens recognized by the sera of patients with oral pemphigoid and understand their effects on normal human buccal mucosa Identifying α6 integrin as a target antigen could improve diagnostic accuracy for oral pemphigoid and help differentiate it from other blistering diseases.

ACIF = anticomplement immunofluorescence; ANA = antinuclear antibody; BMZ = basement membrane zone; C3 = complement component 3; CB = cytoid bodies; DIF = direct immunofluorescence; DG = desquamative gingivitis; Dsg1 = desmoglein 1; Dsg3 = desmoglein 3; H&E = hematoxylin and eosin; HGD = high-grade dysplasia; IIF = indirect immunofluorescence; IgG = immunoglobulin G; IgA = immunoglobulin A; IgM = immunoglobulin M; LE = lupus erythematosus; LGD = low-grade dysplasia; MMP = mucous membrane pemphigoid; MMO = maximum mouth opening; OLDR = oral lichenoid dysplasia; OLL = oral lichenoid lesions; OLP = oral lichen planus; OMLP = oral mucosal lichen planus; OSCC = oral squamous cell carcinoma; OSF = oral submucous fibrosis; PV = pemphigus vulgaris; RAS = recurrent aphthous stomatitis; T1 = tumour size classification (smallest); T4 = tumour size classification (largest); ELISA = enzyme-linked immunosorbent assay; HCV = hepatitis C virus; SCC = squamous cell carcinoma; IF = immunofluorescence.