Conclusions: Data suggest that FUS, TRN1 and TAF15 may participate in a functional pathway in an interdependent way, and imply that the function of TDP-43 may not necessarily be in parallel with, or complementary to, that of FUS, despite each protein sharing many similar structural elements. “
“Research into familial Parkinson’s disease (PD) remained at a virtual standstill in Europe and the US for several decades

until a re-challenge by Japanese selleck products neurologists regarding an autosomal recessive form of PD. In 1965, our research group at Nagoya University examined familial cases of early-onset parkinsonism characterized by autosomal recessive inheritance, diurnal fluctuation of symptoms (alleviation after sleep), foot dystonia, good response to medication, and benign course without dementia. An inborn error of metabolism in some dopamine-related pathway was suspected. The clinical study of four families with the disease, named as “early-onset parkinsonism GSK-3 activation with diurnal fluctuation (EPDF)”, was published in Neurology in 1973. The pathological study of a case in 1993 revealed neuronal loss without Lewy bodies in the substantia nigra. Based on these clinical and pathological evidences, EPDF was defined as a distinct disease entity.

Screening for the EPDF gene was started in 1994 in collaboration with Juntendo University. With the discovery of parkin gene in 1998, EPDF was designated as PARK2. Of our 16 families examined for gene analysis, 15 proved to be PARK2, and the remaining one, PARK6. It was acknowledged long ago that Parkinson’s disease (PD) occurs rarely in familial aggregations. Willige1 collected 12 cases of early-onset parkinsonism and noted a history of familial occurrence in half of them. He proposed regarding

the familial cases as a separate nosological entity under the name of “paralysis agitans juvenilis familialis”, although he failed Ureohydrolase to find essential symptomatic differences from presenile PD. Mjones,2 through a large epidemiological study, indicated a family aggregation. However, in his report there was no mention of clinical manifestations. Research into this sphere remained at a virtual standstill in Europe and the US for several decades thereafter. The re-challenge to familial PD was the discovery by Japanese neurologists of an autosomal recessive form of PD. In 1964, I joined the Neurology Section (Director, Professor I. Sobue), Nagoya University School of Medicine, Nagoya, Japan. In this section, prominent physicians were all working actively and it was full of creative energy. In October 1965, sisters with parkinsonism were admitted to Nagoya University Hospital. I was appointed to these sisters. This was my first and shocking encounter with a novel disease, later known as PARK2. We were interested in their unusual symptoms: diurnal fluctuation or alleviation of difficulties in moving after sleep. We published the cases in Rinsho Shinkeigaku (Tokyo) in 1968.

, 1999; Nishikaku, 2003). The specificity of the immunohistochemical reaction was demonstrated by the absence of staining detected in control tissue slides without the presence

of anti-IFN-γ antibody (Fig. 1a). In omentum tissue sections of uninfected mice, only weak positivity was observed in mononuclear cells (Fig. 1b). After 15 days of Pb18 infection, IFN-γ immunostaining was detected in sparse lymphomononuclear cells at the periphery of omentum granulomas of B10.A susceptible mice (Fig. 1c). In A/J resistant mice, marked positive reaction was found in lymphomononuclear cells at the peripheral foci of necrotic lesions (Fig. 1d), which were mainly observed in this mouse strain. At 120 days post infection, B10.A mice showed disseminated loose lesions with IFN-γ stained cells circumscribing granulomatous foci (Fig. 1e). In A/J mice, https://www.selleckchem.com/products/epz015666.html intense positivity was detected in lymphomononuclear cells forming Gemcitabine several aggregates surrounding central necrosis and compact granulomatous lesions (Fig. 1f). At this later phase of infection, the lesions developed by both mouse

strains showed marked ECM deposition, but with weak immunostaining for IFN-γ (data not shown). After 15 days of infection with the slightly virulent P. brasiliensis isolate Pb265, a similar pattern of IFN-γ staining was detected in both mouse strains when compared with Pb18 inoculated mice at the early stage of infection. Positive lymphomononuclear cells were localized at the periphery of granulomatous lesions (Fig. 2a and b). On the other hand, few IFN-γ positive cells were Dapagliflozin found in the residual lesions of both mouse strains at the later phase of infection with Pb265 (Fig. 2c and d). Figures 3 and 4 show the quantitative analysis of IFN-γ immunohistochemical reaction. The number of immunoreactive cells was similar in the lesions of B10.A and A/J

mice after 15 days postinfection with Pb18. In contrast, the number of IFN-γ positive cells increased in both mouse strains at the later phase of infection with Pb18 (120 days), being significantly higher in A/J mice, when comparing the stage of infection (P < 0.05; 15 vs. 120 days) and also the mouse strain (P < 0.05; B10.A vs. A/J). Regarding the intensity of immunostaining at 15 days post infection with Pb18, the percentage of weakly positive cells predominated over strongly immunostained cells in the lesions of susceptible (68%) and resistant (62%) mice, whereas at 120 days post infection, the number of weakly and strongly immunostained cells was similar in B10.A (55% and 45%, respectively) and in A/J mice (50%). Many immunostained cells were found in B10.A and A/J mice at 15 days post infection with Pb265. The percentage of weakly and strongly positive cells was similar in the susceptible mice (53% and 47%, respectively), but in the resistant ones, there were higher numbers of weakly positive cells (59%).

Two-thirds of patients had coronary disease, one-third had peripheral vascular disease and one quarter had cerebrovascular disease while 70% had some form of vascular disease. An appreciable number of elderly patients (46%) commenced dialysis without permanent access and approximately one-third commenced RRT less than 3 months after nephrologist review. Patients Dorsomorphin solubility dmso on non-dialysis pathways tend to be older,[9, 10] with more functional impairment11 and social isolation[11] but these studies to date are not derived from an Australasian cohort. Elderly ESKD patients who commence

dialysis have considerable mortality. An Australasian study showed 1-year survival of 77%, 2-year survival of 59% and 3-year survival of 45%.[8] Survival of elderly ESKD patients on a non-dialysis pathway is difficult to estimate because of lack of data. Survival without dialysis may be between 9 and 22 months. From ANZDATA and other international registry data, we have accurate information

on the overall survival from the point of MEK inhibitor initiating dialysis within a given age group. It is clear that elderly patients on dialysis have a substantial decrease in actuarial survival compared with the age matched population.[8] The survival of Australasian elderly dialysis patients was as detailed above and was markedly less than the actuarial survival of a similarly aged person not requiring dialysis[12] as shown in Figure 1. These findings have been echoed in publications from other large international registry databases.[1, 13] In a US Renal Data System (USRDS)-based study looking at outcomes of all nursing home residents in the USA following initiation of dialysis, the authors reported mortality rates of 24% in the first 3 months after dialysis initiation and 58% at 12 months.[14] Survival on a non-dialysis pathway is more difficult to determine as there have been few studies, each containing small numbers of patients (Fig.  2). Some studies have reported outcomes on patients of all ages while others have focused on the elderly and the studies

have used different points from which to measure survival, ranging from an epidermal growth factor receptor (eGFR) of 10 or 15 or a putative dialysis date. The reported survival varies between Farnesyltransferase 6 and 23 months in studies with patients of all ages and 9 and 22 months in studies in the elderly. This lack of evidence and variation in mortality makes it difficult for nephrologists to draw conclusions regarding survival on a non-dialysis pathway. Another thing to consider is that the most of these studies were conducted on the UK where practice patterns and characteristics of patients may be different from Australasia. Predictors of survival for elderly patients on dialysis include age, comorbidity score, malnutrition, poor functional status and late referral.

However, it remains to be clarified whether DCs may participate in the pathogenesis of other autoimmune diseases. Previously we have demonstrated that, in primary SS, blood immature myeloid DCs are decreased and mature myeloid DCs are accumulated in salivary glands, suggesting the recruitment of myeloid DCs from blood to inflamed salivary glands. In addition, we demonstrated that numerous IFN-γ-producing CD4+ T cells are also infiltrated into the salivary glands from primary SS patients [2]. Based upon these findings, we proposed a hypothesis that myeloid DCs play a role in pathogenesis of primary SS by initiating Th1 immune response. In this study, we report

that the decrease PD-0332991 chemical structure of blood myeloid DCs and accumulation of salivary gland-infiltrating DCs is universal in the early phase of not only primary SS but also secondary SS, and this alteration was restored spontaneously during the natural clinical course. As shown in Table 1, patients enrolled into this study comprised 24 patients with secondary SS (two men and 22 women, mean age 55·5 years), 29 with primary SS (two men and 27 women, mean age 58·6 years), 11 with SLE (two men and nine women, mean age 25·3 years),

14 with SSc (one man and 13 women, mean age 54·9 years) and 12 with RA (three men and nine women, mean age 55·9 years). In addition, 32 healthy volunteers (12 men and 20 women, mean age 48·0 years) were also enrolled into this study as normal this website controls. All patients presented to our hospital between May 1999 and June 2003 and were diagnosed freshly as having autoimmune diseases. No patients or volunteers had evidence of infections at the time of this study. All patients underwent routine laboratory examinations and

were also examined for a variety of autoantibodies. Informed consent was obtained for this study in accordance Phospholipase D1 with the provisions of the Declaration of Helsinki. All SS patients met the criteria of the Research Committee on SS of the Ministry of Health and Welfare of Japan [12], as well as the European Community criteria [13]. Patients with SLE or SLE-merged secondary SS fulfilled the diagnostic criteria for SLE of the American College of Rheumatology (ACR) [14,15]. Patients with RA or RA-merged secondary SS fulfilled the diagnostic criteria for RA of the ACR [16]. Patients with SSc or SSc-merged secondary SS fulfilled the diagnostic criteria for SSc of the ACR [17]. We determined the onset of SS by a patient complaint about Sicca syndrome in a medical interview (Table 1). In order to assess whether the number of peripheral blood DCs (PBDCs) changes during the natural course of primary SS, six primary SS patients with long-term follow-up were examined sequentially. All the six primary SS patients’ PBDCs were examined in the chronic phase of the disease, 24 months or after the onset of Sicca syndrome [all women, mean age 56·5 years (range 51–71 years)].

It is one of the leading causes of maternal, as well as perinatal morbidity and Dabrafenib in vivo mortality, even in developed countries. Despite intensive research efforts, the aetiology and pathogenesis of pre-eclampsia are not understood completely.

Increasing evidence suggests that an excessive maternal systemic inflammatory response to pregnancy with activation of both the innate and adaptive arms of the immune system is involved in the pathogenesis of the disease [1,2]. We have demonstrated previously that the complement system is activated with increased terminal complex formation in the third trimester of normal human pregnancy, and further in pre-eclampsia, as shown by the elevated amounts of activation markers in the systemic circulation [3]. However, in our recent study, the role of the mannose-binding lectin (MBL)-mediated

lectin pathway has been ruled out in the pathological complement activation observed in pre-eclampsia [4]. Ficolins are pattern recognition molecules of the innate immune system that bind to carbohydrate moieties present on the surface of microbial pathogens, apoptotic and necrotic cells. They act through two distinct routes: by initiating the lectin pathway of complement activation in concert with attached MBL-associated serine proteases (MASPs) and by a primitive opsonophagocytosis [5]. Ficolins are oligomeric proteins consisting of an N-terminal selleck products cysteine-rich region, a collagen-like domain and a C-terminal globular fibrinogen-like domain. The latter is responsible selleck chemical for carbohydrate binding [6]. Three types of ficolins have been identified in humans: ficolin-2 (L-ficolin), ficolin-3 (H-ficolin) and ficolin-1 (M-ficolin). The mRNA of ficolin-2 is expressed primarily

in the liver and its protein product is secreted into the blood circulation. Ficolin-2 exhibits lectin activity toward N-acetyl-glucosamine (GlcNAc) and 1, 3-β-D-glucan. Ficolin-3 mRNA is expressed in the liver and lung. In the liver, ficolin-3 is produced by bile duct epithelial cells and hepatocytes, and is secreted into the bile and circulation. In the lung, ficolin-3 is produced by ciliated bronchial epithelial cells and type II alveolar epithelial cells, and is secreted into the bronchus and alveolus. Ficolin-3 binds to GlcNAc, N-acetyl-galactosamine (GalNAc) and fucose. Ficolin-1 mRNA is expressed in monocytes, the lung and spleen. Its protein product has been identified in secretory granules of neutrophils and monocytes, as well as in type II alveolar epithelial cells. Nevertheless, it is present in the circulation at very low levels compared to ficolin-2 and ficolin-3. Ficolin-1 exhibits binding activity towards GlcNAc, GalNAc and sialic acid [7].

In experiments using influenza virus, autologous B-LCL were infected overnight, whereafter the B-LCL were irradiated and washed selleck kinase inhibitor extensively. After 4–6 days of culture, the allo-specific proliferation of responder T cells was analyzed by flow cytometry. For measurement of suppression on IL-2 production CFSE-labeled D1.50 was cocultured with the indicated M1-specific T-cell

clone at a 1:1 ratio together with autologous B-LCL in the presence of 50 IU/mL IL-2. The Treg clone was prestimulated with 5 μg/mL cognate peptide. After 24 h D1.50 was stimulated with 5 μg/mL cognate peptide, and 1 h later 10 μg/mL Brefeldin A (Sigma-Aldrich) was added. After overnight incubation, the cells were fixed, permeabilized, and stained for CD4 and intracellular IL-2 as described earlier 39. The percentage of IL-2-producing cells was analyzed by flow cytometry. We would like to thank Klara Broadway for technical assistance. The authors declare no conflict of interest. This study was financially supported by a grant from the Netherlands

Organization for Scientific Research (Zon/Mw 917.56.311 to S.H.v.d.B.). Conflict of interest: The authors declare no financial or commercial conflict of interest. “
“Stress activates the hypothalamic-pituitary-adrenocortical axis to promote the release of corticosterone (CORT), which consequently suppresses pathogenic stimulation of the immune system. Paradoxically, however, stress often promotes autoimmunity through yet unknown mechanisms. Opaganib mw Here we investigated how chronic variable stress (CVS), and the associated alterations in CORT levels, affect the susceptibility to experimental autoimmune encephalomyelitis (EAE) in female and male C57BL/6 mice. Under baseline (nonstressed) conditions, females exhibited substantially higher CORT levels and an attenuated EAE with less mortality

than males. However, CVS induced a significantly worsened EAE in females, which Florfenicol was prevented if CORT signaling was blocked. In addition, females under CVS conditions showed a shift toward proinflammatory Th1/Th17 versus Th2 responses and a decreased proportion of CD4+CD25+ Treg cells. This demonstrates that whereas C57BL/6 female mice generally exhibit higher CORT levels and an attenuated form of EAE than males, they become less responsive to the immunosuppressive effects of CORT under chronic stress and thereby prone to a higher risk of destructive autoimmunity. It has been well established that stress may substantially affect the homeostatic regulation of the immune system [1-3]. In most animal models studied thus far, stressful triggers such as fear, maternal deprivation, social threat, or physiological challenge have been shown to induce immunosuppression associated with increased susceptibility to allergies and infectious diseases [1, 4, 5]. These effects are mediated by the hypothalamic-pituitary-adrenal (HPA) axis, a complex network linking the nervous, endocrine and immune systems [6, 7].

In man, hsp90, hsp70, hsp60/Chaperonin and hsp40 families have been characterized.[8] In prokaryotes, GroEL (hsp60) and DnaK (hsp70) are the main hsp families. Stress proteins are ubiquitous and can be detected readily in normal human plasma samples.[9]

Absolute levels of extracellular hsp vary markedly between individuals. For example, reported levels for human plasma hsp60 range between < 1 ng/ml and 1 mg/ml[9] and between 100 pg/ml and 160 ng/ml for CDK inhibitor serum hsp72.[10] Levels of hsp are dynamic during normal physiological activities; exercise increases hsp72 levels in serum by fourfold to eightfold.[11] Therefore, extracellular hsp are continuously present in the circulation of normal individuals and can be increased transiently by several fold without apparent pathology. In addition to functioning as intracellular protein chaperones, hsp modulate the immune system by stimulating both innate and adaptive responses. The term ‘chaperokine’ has been used to describe the dual activity of hsp functioning as both chaperone and cytokine.[12] Once released from a host or pathogen cell, hsp bind to this website cellular receptors to trigger an

innate immune response, including maturation of DC and secretion of pro-inflammatory cytokines and chemokines, for example RANTES (Regulated on Activation Normal T-cell Expressed and Secreted), through Toll-like receptor activation.[13] Processing of cargo proteins carried by hsp occurs, leading to antigen presentation on MHC. Hence hsp link the innate and acquired immune responses to pathogens and have the potential to function as vaccine aminophylline adjuvants in infections and cancer.[14] For

example, hsp70 is an effective and safe adjuvant in neonatal mice and functions effectively via mucosae to generate protective cell-mediated immune responses against herpes simplex virus type-1.[15] Moreover, modified hsp are also capable of inducing cytokine responses. For example, a fusion protein containing Bacillus Calmette–Guérin (BCG)-derived hsp70 and Mycobacterium leprae-derived major membrane protein binds to human DC stimulating production of interleukin-12 p70 through Toll-like receptor 2.[16] Dendritic cells and other cell types possess multiple receptors that bind hsp but the identities and functions of those proposed to modulate the immune system in vivo are not fully understood.[17] The expression profile of these receptors is broad, including, but not limited to, multiple immune, epithelial, endothelial and fibroblast cells and multiple cell types of the central nervous system. Receptors for which evidence supports a role in hsp binding and their distribution on immune cells are shown (Table 2). The relative contribution made by each receptor type to the binding and internalization of hsp by DC is poorly understood.

Figure 4. Overexpression huBCL-2 and huMCL-1 in CD8αα+ iIELs from WT and Il15ra−/− mice Figure S5. Bcl-2 and Bim affect CD8αα+ iIELs survival during in spleen compartment of Il15ra−/− recipients. Figure S6. IL-15-mediated ERK activation in CD8αα+ iIELs is unlikely stimulated by IL-15-induced secreted soluble factor(s) Figure S7. Working model for IL-15-mediated CD8αα+ iIEL survival “
“Diagnostic tests for tuberculosis (TB) using interferon gamma (IFN-γ) responses produced by T lymphocytes after stimulation by early secretory antigen target 6 (ESAT-6), culture filtrate protein see more 10 (CFP-10) or purified protein derivate (PPD) were carried out using ELISA (enzyme-linked immunosorbent assay) in whole blood culture supernatants

from children with suspected TB disease (n = 21), latent TB infection (LTBI; n = 17) and negative controls (NC; n = 21) from Recife, Pernambuco, Brazil. The results were analysed using the ROC (receiver operating characteristic) curves and the areas under the curve (AUC) generated varied from 0.5 to 1.0 with higher values indicating increased discriminatory ability. Comparisons of AUCs were made using non-parametric assumptions, and the

differences were considered significant if P < 0.05. The ROC curve showed a statistical difference (P = 0.015) between the LTBI and NC groups with an AUC of 0.731, TB disease and NC (AUC = 0.780; P = 0.002) PF-6463922 clinical trial and a group with TB (latent infection + disease, n = 38) and NC (AUC = 0.758; P = 0.001) when the antigen used was ESAT-6. No statistical difference was found between the groups when CFP-10 or PPD was used. In conclusion, the ESAT-6 test may be the most appropriate for diagnosis of childhood TB, both LTBI and TB disease, when associated with epidemiological and clinical data, especially in endemic areas such as Brazil. Tuberculosis (TB) is one of the most important infections of humans and a major Glutamate dehydrogenase global public health problem. The World Health Organization (WHO) [1] has annually reported approximately 9.2 million new cases of TB and 1.7 million deaths attributed to this disease. On the other hand, it has been estimated that one-third of

the world population is infected with the intracellular pathogen, Mycobacterium tuberculosis, and one of the most remarkable features of this pathogen is its capacity to generate a latent infection [2, 3]. People that have latent TB infection (LTBI) could be a potential reservoir for future infections, especially when the patient is in childhood and has a compromised immune system [4]. However, depending on the epidemiological situation and the intensity of infection locally, the probability of development of clinical disease after infection with M. tuberculosis may vary [5]. In Brazil, according to the Ministry of Health (2004), 116 000 cases of tuberculosis are reported every year, of which 10% are in children. The country may thus be considered an area where TB is endemic [1].

CD4+ T cells (lanes 1 and 2) and Jurkat cells (lanes 3 and 4). Silver staining of immunoprecipitates, CD4+ T cells (lane 1) and Jurkat cells (lane 3). Immunoprecipitates analysed by Western blotting using anti-FcγRIIIA/B, lane 2 (CD4+ T cells) and lane 4 (Jurkat cells). At 29 kD a broad band with another band at 35 kD was observed in CD4+ T cells and a single band at 29 kD in Jurkat cells. Fig. S7. Immunoprecipitates obtained using anti-FcγRIIIA/B Ku-0059436 ic50 monoclonal antibodies. Proteins stained using silver (left) and blots stained with Coomassie Brilliant Blue R250. Untreated cells (lane 1), terminal complement complex

(TCC) (lane 2), TCC and immune complexes (ICs) (lane 3) and IC-treated cells (lane 4). Arrow point to a protein migrating at approximately 72 kD (Syk). Fig. S8. Inhibition of aggregated human γ-globulin (AHG) binding to CD4+ T cells by see more anti-FcγRIIIA/B monoclonal antibody. 1 × 106 cells treated with AHG-AlexaFluor®488 (5 µg), control cells treated

with isotype antibody (10 µg, left panel) and monoclonal anti-FcγRIIIA/B (10 µg, right panel). “
“Eotaxin-3/CCL26 is an agonist for chemokine receptor 3 (CCR3) and a natural antagonist for CCR1, CCR2 and CCR5. CCL26 expression by non-haematopoietic cells has been well documented; however, no studies to date have demonstrated CCL26 expression by leucocytes. In this study, we investigated the ability of human monocytic cells to produce CCL26 in response to cytokines. We found that interleukin-4 (IL-4) increased the expression of CCL26 messenger RNA (mRNA) and protein in U937 Isotretinoin cells, in human monocytes and in human monocyte-derived macrophages. Tumour necrosis factor-α (TNF-α) and interleukin-1β

(IL-1β) alone did not induce CCL26 expression, yet these pro-inflammatory cytokines synergized with IL-4 to increase CCL26 protein expression. Signal transducer and activator of transcription 6 (STAT6) was not affected by costimulation with TNF-α, suggesting that the synergy between IL-4 and TNF-α occurs at a step downstream of STAT6 activation. Co-incubation of interferon-γ (IFN-γ) with IL-4 had no effect on CCL26 protein release. By contrast, pretreatment with IFN-γ decreased total STAT6 protein, blocked IL-4-mediated STAT6 phosphorylation and decreased IL-4-mediated CCL26 mRNA expression and protein release. These data show that IL-4 and pro-inflammatory cytokines such as TNF-α, IL-1β and IFN-γ regulate CCL26 synthesis in human monocytic cells, which may be important in regulating monocyte inflammatory responses. The eotaxin subfamily of CC chemokines consists of eotaxin/CCL11, eotaxin-2/CCL24 and eotaxin-3/CCL26.1–3 Although they only share 34–39% protein homology, all eotaxins activate cells via CC chemokine receptor 3 (CCR3), which is expressed on several different cell types including eosinophils, basophils, dendritic cells, smooth muscle cells, epithelial cells and fibroblasts (reviewed in Ref. 4). CCL11 and CCL24 are expressed by haematopoietic and non-haematopoietic cells.

Mice (female, 6-week-old, variety BALB/c) were from Research Institute of Animal Production (Velaz, Prague, Czech Republic). The mice had free access to standard pelleted diet and tap water. The animal facilities comply with the European Convention for the Protection of Vertebrate Animals Used for Experimental LY294002 nmr and Other Purposes. The experimental protocol was approved by the Ethics Committee and by the Slovak State Veterinary Committee

of Animal Experimentation. Mice (40 mice per one conjugate) were subcutaneously (sc) primary immunized (1st dose) with conjugate without adjuvant (6 μg oligosaccharide per dose) and subsequently primary sc boosted (2nd dose) without adjuvant 2 weeks after primary injection. Two weeks after primary booster injection, mice were divided

into two groups and were secondary boosted by sc (3rd sc dose) or intraperitoneal (ip, 3rd ip dose) administration of the same conjugate dose without adjuvant. Sera samples were collected at day 14 following each injection. Mice (10 mice in group) three times sc injected with saline in the same time schedule were used as controls. Yeast strain C. albicans CCY 29-3-100 (serotype A) (Culture Collection of Yeasts, Institute of Chemistry, Slovak Academy of Sciences, Slovakia) was cultured on 7% malt extract agar buy Cobimetinib at 28 °C. After 48 h, static cultivation cells were harvested in saline, washed twice with PBS pH 7.4. Viability was specified by flow cytometry with propidium iodide negative staining >99%. Fixation of Candida cells was carried

out very by mixing with 60% ethanol (45:5 v/v) and incubating 15 min at 25 °C, washed twice with PBS and adjusted to 5 × 106 cells/ml with PBS. Ethanol-killed Candida cells were used as control sample in flow cytometric analysis for electronic gates set-up. Levels of induced anti-mannan sera immunoglobulins (IgG, IgM and IgA) were determined by ELISA test, using C. albicans serotype A, C. albicans serotype B or C. guilliermondii mannan in coating step [18]. Antibodies levels were detected at serum dilution 1:100 (n = 10 mice from each group). For the exact expression of IgG, IgM and IgA levels, quantification (in ng/ml) using appropriate calibration curve based on reference mouse serum (Mouse Reference Serum; Bethyl Laboratories, Inc., Montgomery, TX, USA) was done. Statistic analysis was performed using one-way ANOVA test. All data were expressed as mean ± SD. P-values <0.05 were considered statistically significant. Induced C. albicans CCY 29-3-100 (serotype A) whole cell–specific sera immunoglobulin levels (IgG, IgM and IgA, n = 10 mice from each group) were determined by whole cell ELISA test, using C. albicans serotype A cells as yeast and hyphal morphological forms in plate-coating step. The concentrations of coated substances and C.