Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-2

4) in initial binding buffer (1 M (NH)SOin 0.05 M phosphate buffer, pH 6.5). Elution was performed at 0.6 M and 0.4 M (NH)SO. In the Coomassie-stained polyacrylamide gel, one fraction is represented by one lane: lanes 1 – 4 show the flow-through in binding buffer, lanes 5 – 12 the fractions eluted with 0.6 M (NH)SOand lanes 13 – 21 the fractions eluted with 0.4 M (NH)SOand lanes 22 29 represent the fractions eluted in phosphate buffer during the washing step.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-7

different marking techniques (Coomassie-staining, proteins treated with Schiffs reagent and Immuno Blot). 9a: The Coomassie-stained proteins P34, Ovalbumin, BSA are shown (2 μg each lane). 9b: Proteins P34, Ovalbumin, BSA are shown treated with Schiffs reagent (2 μg each lane). 9c: Proteins P34, Ovalbumin, BSA were blotted and immuno-detected with mAb F5.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-5

S were concentrated and equal amounts were loaded on the shown Coomassie-stained gel as described in the Methods section. Lane 0 shows the column feed (soybean feedstock), lanes 1 and 2 the flow-through in binding buffer (0.6 M (NH)SO), lanes 3 and 4 two peaks during the elution step with 0.25 M (NH)SO(comp. Fig. 6) and lane 5 the proteins eluted without (NH)SOin 0.05 M phosphate buffer.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-4

Ed 0.6 M (NH)SO. Protein P34 was eluted using 0.25 M (NH)SO. Subsequently, still bound protein at the column was eluted with phosphate buffer containing no (NH)SO.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-6

In the Methods section. Lanes 1 and 2 represent the flow-through in binding buffer (0.6 M (NH)SO), lanes 3 and 4 the two elution peaks at 0.25 M (NH)SOand lanes 5 and 6 the proteins eluted without (NH)SOin 0.05 M phosphate buffer.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-1

Parated using gel electrophoresis; from each column, two lanes (eluted proteins in binding buffer (flow-through) and at 0.4 M (NH)SO) of every gel were included into this figure. P34 is represented by a band at ~32 kDa. Lane 0 represents the soybean feedstock used as column feed (diluted to 1 μg of total protein), lanes 1 and 2 are eluted fractions collected from Butyl Sepharose 4 Fast Flow, lanes 3 and 4 from Butyl-S Sepharose 6 Fast Flow, lane 5 und 6 from Octyl Sepharose 4 Fast Flow, lane 7 and 8 from Phenyl Sepharose 6 Fast Flow (low sub), lane 9 and 10 from Phenyl Sepharose 6 Fast Flow (high sub) and 11 and 12 from Phenyl Sepharose High Performance.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-8

D on a Coomassie-stained polyacrylamide gel (2 μg each lane): 1 homogenate before centrifugation, 2 oil body after the first centrifugation step, 3 oil body after the last washing step with sodium carbonate, 4 sodium carbonate supernatant, 5 dialysed protein mixture before chromatographic separation.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography-0

D on a Coomassie-stained polyacrylamide gel (2 μg each lane): 1 homogenate before centrifugation, 2 oil body after the first centrifugation step, 3 oil body after the last washing step with sodium carbonate, 4 sodium carbonate supernatant, 5 dialysed protein mixture before chromatographic separation.<p><b>Copyright information:</b></p><p>Taken from "Isolation of soybean protein P34 from oil bodies using hydrophobic interaction chromatography"</p><p>http://www.biomedcentral.com/1472-6750/8/27</p><p>BMC Biotechnology 2008;8():27-27.</p><p>Published online 11 Mar 2008</p><p>PMCID:PMC2292165.</p><p></p

Western blot of tight junction proteins ZO-1 and claudin-3 in IPEC-J2 cells treated with deoxynivalenol (DON).

<p>Cells were grown on inserts and incubated for 24, 48 or 74 hours with DON (0 or 2000 ng/mL) applied from apical or basolateral side in complete medium. ZO-1 (225 kDa) and claudin-3 (22 kDa) expression was analysed by immunoblotting. The housekeeping protein GAPDH (37 kDa) was used as loading control.</p

Cellular distribution of the tight junction protein claudin-3 (CLDN-3) in IPEC-J2 monolayers treated with deoxynivalenol (DON).

<p>Cells were grown on inserts and incubated for 24, 48 or 74 hours either without DON (upper panel) or with 2000 ng/mL DON applied from apical (middle panel) or basolateral side (lower panel) in complete medium. Monolayers were stained for the tight junction associated protein claudin-3 and nuclei stained with DAPI, then detected by immunofluorescence microscopy. All micrographs were taken under identical exposure time and in the centre of each membrane. Micrographs are representative for 3 separate experiments with similar results. Scale bar  =  50 µm.</p