Alterations in pathogen-specific cellular and humoral immunity associated with acute peripheral facial palsy of infectious origin.

BACKGROUND: Peripheral facial palsy (PFP) is a common neurologic symptom which can be triggered by pathogens, autoimmunity, trauma, tumors, cholesteatoma or further local conditions disturbing the peripheral section of the nerve. In general, its cause is often difficult to identify, remaining unknown in over two thirds of cases. As we have previously shown that the quantity and quality of pathogen-specific T cells change during active infections, we hypothesized that such changes may also help to identify the causative pathogen in PFPs of unknown origin. METHODS: In this observational study, pathogen-specific T cells were quantified in blood samples of 55 patients with PFP and 23 healthy controls after stimulation with antigens from varicella-zoster virus (VZV), herpes-simplex viruses (HSV) or borrelia. T cells were further characterized by expression of the inhibitory surface molecule CTLA-4, as well as markers for differentiation (CD27) and proliferation (Ki67). Pathogen-specific antibody responses were analyzed using ELISA. Results were compared with conventional diagnostics. RESULTS: Patients with PFP were more often HSV-seropositive than controls (p = 0.0003), whereas VZV- and borrelia-specific antibodies did not differ between groups. Although the quantity and general phenotypical characteristics of antigen-specific T cells did not differ either, expression of CTLA-4 and Ki67 was highly increased in VZV-specific T cells of 9 PFP patients, of which 5 showed typical signs of cutaneous zoster. In the remaining 4 patients, a causal relationship with VZV was possible but remained unclear by clinical standard diagnostics. A similar CTLA-4- and Ki67-expression profile of borrelia-specific T cells was also found in a patient with acute neuroborreliosis. DISCUSSION: In conclusion, the high prevalence of HSV-seropositivity among PFP-patients may indicate an underestimation of HSV-involvement in PFP, even though HSV-specific T cell characteristics seem insufficient to identify HSV as a causative agent. In contrast, striking alterations in VZV- and borrelia-specific T cell phenotype and function may allow identification of VZV- and borrelia-triggered PFPs. If confirmed in larger studies, antigen-specific immune-phenotyping may have the potential to improve specificity of the clinical diagnosis.

Neurologic Consultations and Headache during Pregnancy and in Puerperium: A Retrospective Chart Review.

Headache is a common symptom during pregnancy and in puerperium that requires careful consideration, as it may be caused by a life-threatening condition. Headaches in pregnant women and women in puerperium are classified as primary or secondary; acute, severe and newly diagnosed headaches should prompt further investigation. We aimed to further characterise the demographic features, symptoms, examination findings, and neuroimaging results of cases of headache during pregnancy and in puerperium. All pregnant women or women in postpartum conditions who attended neurological consultations at the emergency department of the clinic for Gynaecology, Obstetrics and Reproductive Medicine of Saarland University/Germany between 2001/2015 and 2012/2019 were enrolled in this retrospective chart review. Data collected from the charts included demographic/pregnancy characteristics, clinical features and imaging findings. Descriptive statistics as well as binary logistic regression were performed. More than 50% of 97 patients had abnormal findings in their neurological examination. Magnetic resonance imaging findings were pathological for almost 20% of patients-indicating conditions such as cerebral venous thrombosis, reversible posterior leukoencephalopathy, brain tumour and intracranial bleeding. The odds of abnormal neuroimaging results were 2.2-times greater among women with abnormal neurological examination findings than among those with normal examination results. In cases of headache during pregnancy and in puerperium, neuroimaging should be indicated early on. Further research is needed to determine which conditions indicate a need for immediate neuroimaging.

Identification of Neural Mechanisms in First Single-Sweep Analysis in oVEMPs and Novel Normative Data.

BACKGROUND: Bone-conducted (BC) VEMPs provide important tools for measuring otolith function. However, two major drawbacks of this method are encountered in clinical practice-small n10 amplitude and averaging technique. In this study, we present the results of a new VEMP setup measuring technique combined with a novel single-sweep analysis. METHODS: The study included BC oVEMP data from 92 participants for the evaluation of normative data using a novel analysis technique. For evaluating test-retest reliability, the intraclass correlation coefficient (ICC) was used. RESULTS: We found significant n10 amplitude differences in single-sweep analyses after the first and second measurements. Thereby, mathematical analyses of the head movement did not show any differences in the first or second measurements. The normative n10 amplitude was 20.66 µV with an asymmetric ratio (AR) of 7%. The new value of late shift difference (LSD) was 0.01 ms. The test retest-reliability showed good to excellent ICC results in 9 out of 10 measurements. CONCLUSIONS: Our results support a phenomenon in single-sweep analysis of the first stimuli independent of head movement and signal morphology. Furthermore, the values obtained with the new measurement method appear to be more sensitive and may allow an extended diagnostic range due to the new parameter LSD.

Fewer neurocognitive deficits and less brain atrophy by third ventricle measurement in PLWH treated with modern ART: A prospective analysis.

Background: Despite antiretroviral therapy, cognitive dysfunction seems to remain a major issue for people living with human immunodeficiency virus (PLWH). Previous studies showed a correlation between the width of the third ventricle (WTV) and neurocognitive disorders in PLWH. Patients and methods: We investigated prevalence and correlation of neuropsychological disorders using WTV as a brain atrophy marker examined by transcranial sonography and MRI in PLWH and healthy age- and gender-matched controls. We used Becks Depression Inventory (BDI) for depression screening, the questionnaires Fatigue Severity Scale (FSS) for fatigue and Short-Form-36 (SF36) for quality of life (QoL) evaluation and Consortium to establish a registry for Alzheimer's disease (CERAD-PLUS) as neuropsychological test battery. Results: 52 PLWH (47 males) and 28 non-infected controls (23 males) with a median age of 52 years (24-78 years) and 51 years (22-79) were analyzed. WTV correlated significantly with age (p < 0.01) but showed no significantly difference in PLWH (median = 3.4 mm) compared to healthy controls (median = 2.8 mm) (p = 0.085). PLWH had both significantly higher BDI-Scores (p = 0.005) and FSS-Scores (p = 0.012). Controls reported higher QoL (SF-36) with significant differences in most items. However, the overall cognitive performance (CERAD total score) showed no significant difference. The WTV of all subjects correlated with neurocognitive performance measured as CERAD total score (p = 0.009) and trail making tests A (p < 0.001) and B (p = 0.018). There was no correlation between the scores of BDI, FSS, SF-36, and CERAD-PLUS items and WTV. Conclusion: WTV is considered as a predictor of cognitive deficits in neurodegenerative diseases. Nevertheless, we found no significant difference in WTV or overall cognitive performance between PLWH and controls. PLWH suffer more often from depression and fatigue and report reduced QoL when compared to healthy controls.

Case report: cerebral sinus vein thrombosis in two patients with AstraZeneca SARS-CoV-2 vaccination.

SARS-CoV-2 infection is associated with an increased rate of thromboembolic events and mortality. Different vaccines are globally used to limit the pandemic. In this report, we present the case of two young female patients with newly diagnosed cerebral sinus vein thrombosis occurring after injection of the vector-based ChAdOx1 vaccine. Both patients presented with unusual headache only. The two of them used an estrogen-containing contraception, had had a history of deep venous thrombosis, and both had MTHFR mutations. Both patients developed SARS-CoV-2 specific humoral and cellular immunity including both CD4 and CD8 T cells. This rare, but serious complication needs to be considered after vaccination of young females, even if there is no evidence of heparin-induced thrombocytopenia.

Acute stroke treatment and outcome in the oldest old (90 years and older) at a tertiary care medical centre in Germany-a retrospective study showing safety and efficacy in this particular patient population.

BACKGROUND: Stroke is among the most common causes of death and disability worldwide. Despite the relevance of stroke-related disease burden, which is constantly increasing due to the demographic change in industrialized countries with an ageing population and consecutively an increase in age-associated diseases, there is sparse evidence concerning acute stroke treatment and treatment-related outcome in the elderly patient group. This retrospective study aimed at analysing patient characteristics, therapy-related complications and functional outcome in stroke patients aged 90 years or older who underwent acute stroke treatment (i.e. intravenous thrombolysis, mechanical thrombectomy, or both). METHODS: We identified files of all inpatient stays at the Department of Neurology at Saarland University Medical Center (tertiary care level with a comprehensive stroke unit) between June 2011 and December 2018 and filtered for subjects aged 90 years or older at the time of admission. We reviewed patient files for demographic data, symptoms upon admission, (main) diagnoses, comorbidities, and administered therapies. For patients admitted due to acute stroke we reviewed files for therapy-related complications and functional outcome. We compared the modified Rankin scale (mRS) scores upon admission and at discharge for these patients. RESULTS: We identified 566 inpatient stays of subjects aged 90 years or older. Three hundred sixty-seven of the 566 patients (64.8%) were admitted and discharged due to symptoms indicative of stroke. Two hundred eleven patients received a diagnosis of ischaemic stroke. These 211 patients were analysed subsequently. Sixty-four patients qualified for acute stroke treatment (intravenous thrombolysis n = 22, mechanical thrombectomy n = 26, intravenous thrombolysis followed by mechanical thrombectomy n = 16) and showed a significant improvement in their functional status as measured by change in mRS score (admission vs. discharge, p 0.001) with 7 (10.9%) observed potentially therapy-related complications (relevant drop in haemoglobin n = 2, subarachnoidal haemorrhage n = 1, cerebral haemorrhage n = 3, extracranial bleeding n = 1). One intravenous thrombolysis was stopped because of an uncontrollable hypertensive crisis. Patients who did not qualify for these treatments (including those declining acute treatment) did not show a change of their functional status between admission and discharge (p 0.064). CONCLUSION: Our data indicate that acute stroke treatment is effective and safe in the oldest old. Age alone is no criterion to withhold an acute intervention even in oldest old stroke patients.

Short-chain fatty acids and intestinal inflammation in multiple sclerosis: modulation of female susceptibility by microbial products?

BACKGROUND: Multiple Sclerosis (MS) is an autoimmune-mediated disease of the central nervous system. Experimental data suggest a role of intestinal microbiota and microbial products such as short-chain fatty acids (SCFAs) in the pathogenesis of MS. A recent clinical study reported beneficial effects (mediated by immunomodulatory mechanisms) after oral administration of the SCFA propionate in MS patients. Based on available evidence, we investigated whether SCFAs and the fecal inflammation marker calprotectin are altered in MS. METHODS: 76 subjects (41 patients with relapsing-remitting MS and 35 age-matched controls) were investigated in this case-control study. All subjects underwent clinical assessment with established clinical scales and provided fecal samples for a quantitative analysis of fecal SCFA and fecal calprotectin concentrations. Fecal markers were compared between MS patients and controls, and were analyzed for an association with demographic as well as clinical parameters. RESULTS: Median fecal calprotectin concentrations were within normal range in both groups without any group-specific differences. Fecal SCFA concentrations showed a non-significant reduction in MS patients compared to healthy subjects. Female subjects showed significantly reduced SCFA concentrations compared to male subjects. CONCLUSIONS: In our cohort of MS patients, we found no evidence of an active intestinal inflammation. Yet, the vast majority of the investigated MS patients was under immunotherapy which might have affected the outcome measures. The sex-associated difference in fecal SCFA concentrations might at least partially explain female predominance in MS. Large-scale longitudinal studies including drug-naïve MS patients are required to determine the role of SCFAs in MS and to distinguish between disease-immanent effects and those caused by the therapeutic regime.

Consecutive Eyeball Pressure Tests Reflect Clinically Relevant Vagal Dysfunction and Recovery in a Patient With Guillain-Barré-Syndrome With Tenacious Cardiac Dysautonomia.

Cardiac dysautonomia is a potentially life-threatening complication of Guillain-Barré syndrome (GBS). Proper and prompt recognition of patients at risk and subsequent intensive care unit (ICU) monitoring are mandatory to prevent fatal outcome. Eyeball pressure testing (EP) has been suggested as an easy applicable bedside test for vagal overreactivity in GBS and thus identifying patients at risk. Yet, there is only sparse follow-up data concerning the course of EP findings in GBS. We report a 25 years-old male patient with GBS who underwent consecutive EP (n = 11) during his ICU stay over a period of 11 weeks. The series of tests performed in this patient (and corresponding clinical events) show that EP data might represent an approximation of vagal dysfunction and vagal recovery in GBS. Interestingly, we observed a much longer duration of pathological EP compared to a previous report. The tenacious cardiac dysautonomia in this patient necessitated long-term application of a transvenous temporary pacemaker.

Effects of a Feedback-Demanding Stroke Clock on Acute Stroke Management: A Randomized Study.

BACKGROUND AND PURPOSE: This randomized study aimed to evaluate whether the use of a stroke clock demanding active feedback from the stroke physician accelerates acute stroke management. METHODS: For this randomized controlled study, a large-display alarm clock was installed in the computed tomography room, where admission, diagnostic work-up, and intravenous thrombolysis occurred. Alarms were set at the following target times after admission: (1) 15 minutes (neurological examination completed); (2) 25 minutes (computed tomography scanning and international normalized ratio determination by point-of-care laboratory completed); and (3) 30 minutes (intravenous thrombolysis started). The responsible stroke physician had to actively provide feedback by pressing a buzzer button. The alarm could be avoided by pressing the button before time out. Times to therapy decision (primary end point, defined as the end of all diagnostic work-up required for decision for or against recanalizing treatment), neurological examination, imaging, point-of-care laboratory, needle, and groin puncture were assessed by a neutral observer. Functional outcome (modified Rankin Scale) was assessed at day 90. RESULTS: Of 107 participants, 51 stroke clock patients exhibited better stroke-management metrics than 56 control patients. Times from door to (1) end of all indicated diagnostic work-up (treatment decision time; 16.73 versus 26.00 minutes, P<0.001), (2) end of neurological examination (7.28 versus 10.00 minutes, P<0.001), (3) end of computed tomography (11.17 versus 14.00 minutes, P=0.002), (4) end of computed tomography angiography (14.00 versus 17.17 minutes, P=0.001), (5) end of point-of-care laboratory testing (12.14 versus 20.00 minutes, P<0.001), and (6) needle times (18.83 versus 47.00 minutes, P=0.016) were improved. In contrast, door-to-groin puncture times and functional outcomes at day 90 were not significantly different. CONCLUSIONS: This study showed that the use of a stroke clock demanding active feedback significantly improves acute stroke-management metrics and, thus, represents a potential low-cost strategy for streamlining time-sensitive stroke treatment.

[Influence of the "distance to thrombus" in acute middle cerebral artery occlusion : A predictor for the clinical outcome after endovascular treatment]. / Einfluss der Distanz zum Thrombus bei akutem Verschluss der Arteria cerebi media : Ein Prädiktor für das klinische Outcome nach mechanischer Thrombektomie.

BACKGROUND: The therapy strategy of patients with acute stroke of the middle cerebral artery (MCA) is influenced by the location of the occlusion. The purpose of this study was to analyze the clinical outcome in patients with acute ischemic MCA occlusion according to the location of occlusion who underwent endovascular treatment (EVT). METHODS: A total of 54 patients (age 73 ± 15 years; 59% female) with acute ACM occlusion treated with EVT were included. In coronary reformatted CT angiography images, the distance to the thrombus (DT), i.e. the distance from the carotid T to the beginning of the thrombus, was measured. Correlations between DT, clinical symptoms, and clinical outcome of patients who underwent EVT were analyzed. RESULTS: DT correlated with clinical symptoms measured by the National Institutes of Health Stroke Scale (NIHSS; p = 0.017; R = -0.324) at baseline. DT also correlated with the modified Rankin scale after 90 days (90-day mRS; p = 0.014; R = -0.333). DT was a predictor for a good clinical outcome (mRS after 90 days) after EVT; odds ratio 1.113 (P = 0.02; 95% confidence interval [CI] 1.017-1.219). A DT >10 mm correlated significantly (p = 0.036) with a good clinical outcome (90-day mRS ≤2). CONCLUSION: DT correlates with the clinical symptoms of patients with acute MCA occlusion. In addition, DT is an independent predictor of the clinical outcome of patients suffering from acute stroke due to MCA occlusion.

Sonography of optic nerve sheath diameter identifies patients with middle cerebral artery infarction at risk of a malignant course: a pilot prospective observational study.

INTRODUCTION: To assess the value of optic nerve sheath diameter (ONSD) measurements at different time points to predict the malignant evolution in middle cerebral artery (MCA) infarction and to investigate the relationship between ONSD and infarct volume on follow-up computed tomography (CT). METHODS: In a single-center prospective observational study, we recruited patients with MCA infarction and age- and sex-matched controls. Clinical characteristics including NationaI Institutes of Health Stroke Scale (NIHSS) and ONSD measurement were assessed during the first five days after symptom onset. Volumetric analysis of the infarction was performed by a neuroradiologist, who was blinded to results of ONSD measurement and clinical examinations, based on  CT scans. RESULTS: We enrolled 29 patients with MCA infarction, including 10 with malignant MCA (mMCA) infarction and 14 controls. Mean ONSD on admission was already larger in patients who had developed an mMCA (5.99 ± 0.32 mm) compared to patients with MCA infarction (4.98 ± 0.53 mm; P = 0.003), and to control patients (4.57 ± 0.29 mm; P < 0.001). Correlation was observed between the ONSD mean value bilateral measures per individual and volumetric evaluation of cerebral infarction in the CT scan after one day (r = 0.623; P = 0.002). An ONSD value of 5.6 mm predicted an mMCA with a sensitivity of 100% and specificity of 90% yielding a positive predictive value of 83% and negative predictive value of 100%. CONCLUSIONS: ONSD measurement might be accurate for the noninvasive detection of increased ICP and for the recognition of patients being likely to develop mMCA.

Intravenous thrombolysis with tPA and cortical involvement increase the risk of early poststroke seizures: Results of a case-control study.

The aim of this study was to identify the risk factors for early poststroke seizures (PSS) in patients with acute ischemic stroke. We undertook a case-control study at a single stroke center. Patients with seizure occurring during the first 7 days following ischemic stroke admitted between 2010 and 2016 were retrospectively identified and matched with controls (patients with stroke without early PSS) for age and sex. We included 79 cases and 158 controls. Blood sugar levels on admission, stroke localization, National Institutes of Health Stroke Scale (NIHSS) and Rankin score, and intravenous (i.v.) thrombolysis with recombinant tissue plasminogen activator (rtPA) were statistically associated with early PSS in univariate analysis. Multiple logistic regression after forward and backward variable selection identified cortical stroke localization (odds ratio (OR): 2.49; 95% confidence intervals (CI): 1.35 to 4.59; p = 0.003) and i.v. thrombolysis (OR: 2.26; 95% CI: 1.16 to 4.43; p = 0.008) as variables independently associated with early PSS. Cortical involvement and i.v. thrombolysis are independent risk factors associated with the occurrence of early PSS. This association is not explained by age or sex, concomitant drugs, diabetes or alcoholism, sodium and cholesterol levels, blood pressure on admission, stroke etiology or severity, and hemorrhage following i.v. thrombolysis. Further studies are required to fully elucidate the association between different reperfusion therapies and early PSS. This article is part of the Special Issue "Seizures & Stroke".

Primary HIV infection presenting with Kaposi sarcoma and limbic encephalitis.

The development of anti-N-methyl-D-aspartate (NMDA) receptor encephalitis is often associated with neoplasia or infectious diseases as antibodies against neurons or synaptic proteins surface. A 30-year-old male patient was admitted to our department because of neurocognitive symptoms, particularly memory difficulties which had appeared a year prior and since then had been increasing. He had a medical history of smoking and hypertension. On examination, there were no focal neurological deficits. However, neuropsychological tests confirmed a lack of concentration and short-term memory impairment. Brain magnetic resonance imaging (MRI) and electroencephalography (EEG) remained unremarkable. Cerebrospinal fluid (CSF) analysis revealed a low lymphocytic pleocytosis without oligoclonal bands. Serum testing for human immunodeficiency virus (HIV) was positive with 420,000 HIV-1-RNA copies/ml. On a more detailed physical examination, a large number of purple patches were found on the entire body, which a biopsy confirmed to be Kaposi sarcoma (KS). A positive serum and CSF NMDA receptor antibody titer (serum 1:280; CSF 1:8) confirmed the diagnosis of an AIDS-associated anti-NMDA receptor encephalitis; therefore, we treated him with antiretroviral and immunosuppressive therapy. After 12 months, the KS lesions faded and the cognitive deficits improved slightly. Our case highlights that a detailed clinical examination and searching for neoplasia and/or an infection are helpful, though often neglected, tools for detecting an anti-NMDA receptor encephalitis.

Prehospital Stroke Management Optimized by Use of Clinical Scoring vs Mobile Stroke Unit for Triage of Patients With Stroke: A Randomized Clinical Trial.

Importance: Transferring patients with large-vessel occlusion (LVO) or intracranial hemorrhage (ICH) to hospitals not providing interventional treatment options is an unresolved medical problem. Objective: To determine how optimized prehospital management (OPM) based on use of the Los Angeles Motor Scale (LAMS) compares with management in a Mobile Stroke Unit (MSU) in accurately triaging patients to the appropriate hospital with (comprehensive stroke center [CSC]) or without (primary stroke center [PSC]) interventional treatment. Design, Setting, and Participants: In this randomized multicenter trial with 3-month follow-up, patients were assigned week-wise to one of the pathways between June 15, 2015, and November 15, 2017, in 2 regions of Saarland, Germany; 708 of 824 suspected stroke patients did not meet inclusion criteria, resulting in a study population of 116 adult patients. Interventions: Patients received either OPM based on a standard operating procedure that included the use of the LAMS (cut point ≥4) or management in an MSU (an ambulance with vascular imaging, point-of-care laboratory, and telecommunication capabilities). Main Outcomes and Measures: The primary end point was the proportion of patients accurately triaged to either CSCs (LVO, ICH) or PSCs (others). Results: A predefined interim analysis was performed after 116 patients of the planned 232 patients had been enrolled. Of these, 53 were included in the OPM group (67.9% women; mean [SD] age, 74 [11] years) and 63 in the MSU group (57.1% women; mean [SD] age, 75 [11] years). The primary end point, an accurate triage decision, was reached for 37 of 53 patients (69.8%) in the OPM group and for 63 of 63 patients (100%) in the MSU group (difference, 30.2%; 95% CI, 17.8%-42.5%; P < .001). Whereas 7 of 17 OPM patients (41.2%) with LVO or ICH required secondary transfers from a PSC to a CSC, none of the 11 MSU patients (0%) required such transfers (difference, 41.2%; 95% CI, 17.8%-64.6%; P = .02). The LAMS at a cut point of 4 or higher led to an accurate diagnosis of LVO or ICH for 13 of 17 patients (76.5%; 6 triaged to a CSC) and of LVO selectively for 7 of 9 patients (77.8%; 2 triaged to a CSC). Stroke management metrics were better in the MSU group, although patient outcomes were not significantly different. Conclusions and Relevance: Whereas prehospital management optimized by LAMS allows accurate triage decisions for approximately 70% of patients, MSU-based management enables accurate triage decisions for 100%. Depending on the specific health care environment considered, both approaches are potentially valuable in triaging stroke patients. Trial Registration: ClinicalTrials.gov identifier: NCT02465346.

Mobile Stroke Units - Cost-Effective or Just an Expensive Hype?

PURPOSE OF REVIEW: Acute stroke is a treatable disease. Nevertheless, only a minority of patients obtain guideline-adjusted therapy. One major reason is the small time window in which therapies have to be administered in order to reverse or mitigate brain injury and prevent disability. The Mobile Stroke Unit (MSU) concept, available for a decade now, is spreading worldwide, comprising ambulances, fully equipped with computed tomography, laboratory unit and telemedicine connection to the stroke centre and staffed with a specialised stroke team. Besides its benefits, this concept adds a relevant amount of costs to health services. RECENT FINDINGS: The feasibility of the MSU and its impact on reducing treatment times have been proven by several research trials. In addition, pre-hospital stroke diagnosis including computed tomographic angiography analysis facilitates correct triage of patients, needing mechanical recanalization, thereby reducing the number of secondary or inter-hospital transfers. Even so, the concept is not yet fully implemented on a broad scale. One reason is the still open question of cost-effectiveness. There are assumptions based on the randomised trials of MSUs hinting towards an acceptable amount of money per quality-adjusted life years and overall cost-effectiveness. Up to now, neither a prospective analysis nor a consideration of secondary transfer avoidance is available. The MSU concept is an innovative and impactful strategy to improve stroke management, especially in times of constraints in healthcare economics and health care professionals. Prospective information is needed to answer the cost-effectiveness question satisfactorily.

Assay for improved detection of antigen-specific immune cells from extrasanguinous fluids.

In this approach, pre-stained cells from extrasanguinous fluids (ESFs) are stimulated in the presence of blood from the same individual. Thus, blood-derived antigen-presenting cells enable stimulation of both ESF- and blood T cells. Pre-staining allows distinction of T cells from ESF and blood, and simultaneous analysis of antigen-specific T cells in both compartments.