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Merck & Co silica gel thin layer chromatography (tlc) plates
Silica Gel Thin Layer Chromatography (Tlc) Plates, supplied by Merck & Co, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Becton Dickinson mouse anti-btk monoclonal antibody
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Millipore mf 0.45-μm ha membrane filters
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SeraCare Life Sciences fluorescein isothiocyanate (fitc)-labelled goat anti-human immunoglobulin igg (h + l)
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System Biosciences Inc exoab antibody kit for hsp70
Comparison of AFSC-EV protein content and EV markers. ( a ) Protein quantification of AFSC-EV preparations using the Pierce Bradford assay. Data are shown as mean ± SD n = 3. No difference was found between preparations isolated with UC, ExoQuick and TEIR. AFSC-EV preparations isolated using qEV had lower protein content than those isolated using Exo-PREP (p < 0.05). ( b ) Correlation analysis between total number of particles analyzed with Nanoparticle Tracking Analysis (Fig. ) and EV protein concentration (μg/μL) in each preparation of AFSC-EVs obtained by different isolation techniques [p = 0.25, r = 0.6 (95% CI −0.56 to 0.97)]. ( c ) Expression of canonical EV markers <t>Hsp70,</t> CD63, Flotillin-1, and TSG101 obtained by Western blot analysis for the different isolation techniques. All AFSC-EV isolation techniques showed no evidence of residual cellular debris, as evidenced by a lack of H3K27me3 protein expression. AFSCs (parent cells) and AFSC-conditioned medium (AFSC-CM; the initial starting material from which all techniques were derived), are shown as positive controls. Representative photo from n = 3 replicate analyses.
Exoab Antibody Kit For Hsp70, supplied by System Biosciences Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Millipore silane-preptm slides
Comparison of AFSC-EV protein content and EV markers. ( a ) Protein quantification of AFSC-EV preparations using the Pierce Bradford assay. Data are shown as mean ± SD n = 3. No difference was found between preparations isolated with UC, ExoQuick and TEIR. AFSC-EV preparations isolated using qEV had lower protein content than those isolated using Exo-PREP (p < 0.05). ( b ) Correlation analysis between total number of particles analyzed with Nanoparticle Tracking Analysis (Fig. ) and EV protein concentration (μg/μL) in each preparation of AFSC-EVs obtained by different isolation techniques [p = 0.25, r = 0.6 (95% CI −0.56 to 0.97)]. ( c ) Expression of canonical EV markers <t>Hsp70,</t> CD63, Flotillin-1, and TSG101 obtained by Western blot analysis for the different isolation techniques. All AFSC-EV isolation techniques showed no evidence of residual cellular debris, as evidenced by a lack of H3K27me3 protein expression. AFSCs (parent cells) and AFSC-conditioned medium (AFSC-CM; the initial starting material from which all techniques were derived), are shown as positive controls. Representative photo from n = 3 replicate analyses.
Silane Preptm Slides, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Miles Scientific 10 × 20 cm hptlc-ghlf 3 normal phase silica tlc plate
Comparison of AFSC-EV protein content and EV markers. ( a ) Protein quantification of AFSC-EV preparations using the Pierce Bradford assay. Data are shown as mean ± SD n = 3. No difference was found between preparations isolated with UC, ExoQuick and TEIR. AFSC-EV preparations isolated using qEV had lower protein content than those isolated using Exo-PREP (p < 0.05). ( b ) Correlation analysis between total number of particles analyzed with Nanoparticle Tracking Analysis (Fig. ) and EV protein concentration (μg/μL) in each preparation of AFSC-EVs obtained by different isolation techniques [p = 0.25, r = 0.6 (95% CI −0.56 to 0.97)]. ( c ) Expression of canonical EV markers <t>Hsp70,</t> CD63, Flotillin-1, and TSG101 obtained by Western blot analysis for the different isolation techniques. All AFSC-EV isolation techniques showed no evidence of residual cellular debris, as evidenced by a lack of H3K27me3 protein expression. AFSCs (parent cells) and AFSC-conditioned medium (AFSC-CM; the initial starting material from which all techniques were derived), are shown as positive controls. Representative photo from n = 3 replicate analyses.
10 × 20 Cm Hptlc Ghlf 3 Normal Phase Silica Tlc Plate, supplied by Miles Scientific, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Comparison of AFSC-EV protein content and EV markers. ( a ) Protein quantification of AFSC-EV preparations using the Pierce Bradford assay. Data are shown as mean ± SD n = 3. No difference was found between preparations isolated with UC, ExoQuick and TEIR. AFSC-EV preparations isolated using qEV had lower protein content than those isolated using Exo-PREP (p < 0.05). ( b ) Correlation analysis between total number of particles analyzed with Nanoparticle Tracking Analysis (Fig. ) and EV protein concentration (μg/μL) in each preparation of AFSC-EVs obtained by different isolation techniques [p = 0.25, r = 0.6 (95% CI −0.56 to 0.97)]. ( c ) Expression of canonical EV markers Hsp70, CD63, Flotillin-1, and TSG101 obtained by Western blot analysis for the different isolation techniques. All AFSC-EV isolation techniques showed no evidence of residual cellular debris, as evidenced by a lack of H3K27me3 protein expression. AFSCs (parent cells) and AFSC-conditioned medium (AFSC-CM; the initial starting material from which all techniques were derived), are shown as positive controls. Representative photo from n = 3 replicate analyses.

Journal: Scientific Reports

Article Title: The Regenerative Potential of Amniotic Fluid Stem Cell Extracellular Vesicles: Lessons Learned by Comparing Different Isolation Techniques

doi: 10.1038/s41598-018-38320-w

Figure Lengend Snippet: Comparison of AFSC-EV protein content and EV markers. ( a ) Protein quantification of AFSC-EV preparations using the Pierce Bradford assay. Data are shown as mean ± SD n = 3. No difference was found between preparations isolated with UC, ExoQuick and TEIR. AFSC-EV preparations isolated using qEV had lower protein content than those isolated using Exo-PREP (p < 0.05). ( b ) Correlation analysis between total number of particles analyzed with Nanoparticle Tracking Analysis (Fig. ) and EV protein concentration (μg/μL) in each preparation of AFSC-EVs obtained by different isolation techniques [p = 0.25, r = 0.6 (95% CI −0.56 to 0.97)]. ( c ) Expression of canonical EV markers Hsp70, CD63, Flotillin-1, and TSG101 obtained by Western blot analysis for the different isolation techniques. All AFSC-EV isolation techniques showed no evidence of residual cellular debris, as evidenced by a lack of H3K27me3 protein expression. AFSCs (parent cells) and AFSC-conditioned medium (AFSC-CM; the initial starting material from which all techniques were derived), are shown as positive controls. Representative photo from n = 3 replicate analyses.

Article Snippet: Hsp70, CD63, Flotillin-1, and TSG101 were measured from AFSC-EVs, and from 20 μg total protein from AFSC-CM, and AFSCs using ExoAb Antibody Kit for Hsp70 (System Biosciences, Palo Alto, CA; primary antibody: rabbit anti-human, 1:1,000 dilution; secondary antibody: goat anti-rabbit HRP, 1:10,000 dilution), CD63 (System Biosciences, Palo Alto, CA; primary antibody: rabbit anti-human, 1:1,000 dilution; secondary antibody: goat anti-rabbit HRP, 1:10,000 dilution), TSG101 (Santa Cruz Biotechnology, Dallas, TX; primary antibody mouse anti-rat, 1:500 dilution; secondary antibody: goat anti-mouse HRP, 1:3,000 dilution), and Flotillin-1 (BD Transduction Laboratories, San Jose, CA; primary antibody mouse anti-rat, 1:1,000 dilution; secondary antibody: goat anti-mouse HRP, 1:3,000 dilution).

Techniques: Comparison, Bradford Assay, Isolation, Protein Concentration, Expressing, Western Blot, Derivative Assay