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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV130200	1/2	Homo sapiens	BALF	IAF	Levänen B	2013	17%
Study summary Full title Altered microRNA profiles in bronchoalveolar lavage fluid exosomes in asthmatic patients All authors Levänen B, Bhakta NR, Torregrosa Paredes P, Barbeau R, Hiltbrunner S, Pollack JL, Sköld CM, Svartengren M, Grunewald J, Gabrielsson S, Eklund A, Larsson BM, Woodruff PG, Erle DJ, Wheelock ÅM Journal J Allergy Clin Immunol Abstract BACKGROUND: Asthma is characterized by increased airway narrowing in response to nonspecific stimuli (show more...)BACKGROUND: Asthma is characterized by increased airway narrowing in response to nonspecific stimuli. The disorder is influenced by both environmental and genetic factors. Exosomes are nanosized vesicles of endosomal origin released from inflammatory and epithelial cells that have been implicated in asthma. In this study we characterized the microRNA (miRNA) content of exosomes in healthy control subjects and patients with mild intermittent asthma both at unprovoked baseline and in response to environmental challenge. OBJECTIVE: To investigate alterations in bronchoalveolar lavage fluid (BALF) exosomal miRNA profiles due to asthma, and following subway air exposure. METHODS: Exosomes were isolated from BALF from healthy control subjects (n = 10) and patients with mild intermittent asthma (n = 10) after subway and control exposures. Exosomal RNA was analyzed by using microarrays containing probes for 894 human miRNAs, and selected findings were validated with quantitative RT-PCR. Results were analyzed by using multivariate modeling. RESULTS: The presence of miRNAs was confirmed in exosomes from BALF of both asthmatic patients and healthy control subjects. Significant differences in BALF exosomal miRNA was detected for 24 miRNAs with a subset of 16 miRNAs, including members of the let-7 and miRNA-200 families, providing robust classification of patients with mild nonsymptomatic asthma from healthy subjects with 72% cross-validated predictive power (Q(2) = 0.72). In contrast, subway exposure did not cause any significant alterations in miRNA profiles. CONCLUSION: These studies demonstrate substantial differences in exosomal miRNA profiles between healthy subjects and patients with unprovoked, mild, stable asthma. These changes might be important in the inflammatory response leading to bronchial hyperresponsiveness and asthma. (hide) EV-METRIC 17% (67th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type BALF Sample origin NAY Focus vesicles exosomes Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture IAF Protein markers EV: CD63/ MHC2 non-EV: Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type BALF Separation Method Immunoaffinity capture Selected surface protein(s) MHC2 Western Blot Antibody details provided? No Detected EV-associated proteins MHC2 ELISA Antibody details provided? No Detected EV-associated proteins MHC2

	EV130200	2/2	Homo sapiens	BALF	(d)(U)C Filtration	Levänen B	2013	14%
Study summary Full title Altered microRNA profiles in bronchoalveolar lavage fluid exosomes in asthmatic patients All authors Levänen B, Bhakta NR, Torregrosa Paredes P, Barbeau R, Hiltbrunner S, Pollack JL, Sköld CM, Svartengren M, Grunewald J, Gabrielsson S, Eklund A, Larsson BM, Woodruff PG, Erle DJ, Wheelock ÅM Journal J Allergy Clin Immunol Abstract BACKGROUND: Asthma is characterized by increased airway narrowing in response to nonspecific stimuli (show more...)BACKGROUND: Asthma is characterized by increased airway narrowing in response to nonspecific stimuli. The disorder is influenced by both environmental and genetic factors. Exosomes are nanosized vesicles of endosomal origin released from inflammatory and epithelial cells that have been implicated in asthma. In this study we characterized the microRNA (miRNA) content of exosomes in healthy control subjects and patients with mild intermittent asthma both at unprovoked baseline and in response to environmental challenge. OBJECTIVE: To investigate alterations in bronchoalveolar lavage fluid (BALF) exosomal miRNA profiles due to asthma, and following subway air exposure. METHODS: Exosomes were isolated from BALF from healthy control subjects (n = 10) and patients with mild intermittent asthma (n = 10) after subway and control exposures. Exosomal RNA was analyzed by using microarrays containing probes for 894 human miRNAs, and selected findings were validated with quantitative RT-PCR. Results were analyzed by using multivariate modeling. RESULTS: The presence of miRNAs was confirmed in exosomes from BALF of both asthmatic patients and healthy control subjects. Significant differences in BALF exosomal miRNA was detected for 24 miRNAs with a subset of 16 miRNAs, including members of the let-7 and miRNA-200 families, providing robust classification of patients with mild nonsymptomatic asthma from healthy subjects with 72% cross-validated predictive power (Q(2) = 0.72). In contrast, subway exposure did not cause any significant alterations in miRNA profiles. CONCLUSION: These studies demonstrate substantial differences in exosomal miRNA profiles between healthy subjects and patients with unprovoked, mild, stable asthma. These changes might be important in the inflammatory response leading to bronchial hyperresponsiveness and asthma. (hide) EV-METRIC 14% (57th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type BALF Sample origin NAY Focus vesicles exosomes Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Filtration Adj. k-factor 149.8 (pelleting) Protein markers EV: non-EV: Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type BALF Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 120 Pelleting: rotor type 45Ti Pelleting: adjusted k-factor 149.8 Filtration steps 0.22µm or 0.2µm

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EV-TRACK ID	EV130200
species	Homo sapiens
sample type	BALF
condition	NAY
separation protocol	IAF	(d)(U)C Filtration
Exp. nr.	1	2
EV-METRIC %	17	14