title: Immune profiling of plasma and cervical secretions using recycling immunoaffinity chromatography.
abstract: Small volumes of cervical secretions have limited measurements of immunity at the cervix, which may be important to studies of human papillomavirus (HPV). We report the use of recycling immunoaffinity chromatography to efficiently study immune profiles in cervical secretions. Frozen pairs of plasma and cervical secretions (collected on ophthalmic sponges) were selected randomly from women with normal cervical cytology (n = 50) participating in a natural history study of HPV in Guanacaste, Costa Rica. Single 25- micro l aliquots of plasma and (diluted) cervical secretions were assayed for interleukin (IL) -1 beta, -2, -4, -6, -8, -10, -12, -13, -15, IFN-alpha, -beta, -gamma, tumor necrosis factor-alpha, -beta, RANTES (regulated on activation normal T-cell express and secreted), MCP-1 (monocyte chemoattractant protein), -2, -3, macrophage inflammatory protein-1 alpha, -1 beta (regulated on activation normal T-cell express and secreted), macrophage colony-stimulating factor, IgG, IgA, and cyclooxygenase 2. All of the specimens were tested as blind replicates, and refrozen plasma was retested 4 months later. To evaluate the reproducibility of the repeat measurements and to examine the correlation between plasma and cervical secretions, we calculated kappa values with 95% confidence intervals among categorized analyte values and Spearman correlation coefficients (rho) among detectable, continuous analyte values. Measurements of all of the analytes in either plasma or cervical secretions were highly reproducible, with all of the kappa > or = 0.78 (70% above 0.90), and all of the rho > or = 0.88 (96% above 0.90). Only IL-1 beta (kappa = 0.60 and rho = 0.82) and IL-6 (kappa = 0.50 and rho = 0.78) levels were strongly correlated between plasma and cervical secretions. IFN-gamma, tumor necrosis factor-beta, RANTES, MCP-1, MCP -2, macrophage inflammatory protein-1 alpha, and macrophage colony-stimulating factor levels were especially poorly correlated between plasma and cervical secretions (kappa < or = 0.25 and rho < or = 0.25). We conclude that recycling immunoaffinity chromatography is a reproducible method of measuring immune profiles from biological specimens, and immune profiles are not well correlated between plasma and cervical secretions, perhaps necessitating cervical collections to study cervix-specific immunity in HPV natural history studies.
authors.affiliation: Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Department of Health and Human Services, Bethesda, Maryland 20892-7234, USA. castlep@mail.nih.gov
authors.forename: Philip E
authors.initials: PE
authors.lastname: Castle
authors.forename: Terry M
authors.initials: TM
authors.lastname: Phillips
authors.forename: Allan
authors.initials: A
authors.lastname: Hildesheim
authors.forename: Rolando
authors.initials: R
authors.lastname: Herrero
authors.forename: M Concepcion
authors.initials: MC
authors.lastname: Bratti
authors.forename: Ana Cecilia
authors.initials: AC
authors.lastname: Rodríguez
authors.forename: Lidia Ana
authors.initials: LA
authors.lastname: Morera
authors.forename: Ruth
authors.initials: R
authors.lastname: Pfeiffer
authors.forename: Martha L
authors.initials: ML
authors.lastname: Hutchinson
authors.forename: Ligia A
authors.initials: LA
authors.lastname: Pinto
authors.forename: Mark
authors.initials: M
authors.lastname: Schiffman
mesh_terms: D000328:Adult; D017677:Age Distribution; D000368:Aged; D015415:Biomarkers / Q000097:blood*; D002582:Cervix Mucus / Q000821:virology; D018932:Chemokine CCL2 / Q000032:analysis; D018946:Chemokine CCL5 / Q000032:analysis; D054412:Chemokine CCL8; D002845:Chromatography / Q000379:methods*; D015331:Cohort Studies; D016001:Confidence Intervals; D003364:Costa Rica / Q000453:epidemiology; D005260:Female; D006801:Humans; D007158:Immunologic Techniques; D015994:Incidence; D008403:Mass Screening / Q000379:methods; D008875:Middle Aged; D018945:Monocyte Chemoattractant Proteins / Q000032:analysis; D027383:Papillomaviridae / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000175:diagnosis* / Q000453:epidemiology*; D011336:Probability; D012307:Risk Factors; D013048:Specimen Handling; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis
chemical_list: D015415:Biomarkers; C515453:CCL8 protein, human; D018932:Chemokine CCL2; D018946:Chemokine CCL5; D054412:Chemokine CCL8; D018945:Monocyte Chemoattractant Proteins; D014409:Tumor Necrosis Factor-alpha