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Boehringer Mannheim dispase solution
Quantitative measurements of keratinocyte cell adhesion as determined by a novel <t>dispase-based</t> assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.
Dispase Solution, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 92/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Tyrosine Phosphorylation and Src Family Kinases Control Keratinocyte Cell-Cell Adhesion "

Article Title: Tyrosine Phosphorylation and Src Family Kinases Control Keratinocyte Cell-Cell Adhesion

Journal: The Journal of Cell Biology

doi:

Quantitative measurements of keratinocyte cell adhesion as determined by a novel dispase-based assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.
Figure Legend Snippet: Quantitative measurements of keratinocyte cell adhesion as determined by a novel dispase-based assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.

Techniques Used: Incubation, Infection, Expressing

Decreased strength of calcium-induced cell adhesion as a consequence of tyrosine kinase inhibition or lack of the Fyn kinase. ( A ) Primary keratinocytes under growing conditions were either tested as untreated controls ( Ctr ) or pretreated for 2 h with Genistein. Incubation was then continued for additional 24 h under high calcium conditions. Cultures were examined as such ( left panels ) or after dispase treatment for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the tyrosine kinase inhibitor-treated cultures. No such areas were evident in control cultures even after prolonged dispase exposure ( > 30 min). Instead, control cells eventually detached from the dish as a confluent sheet. Similar results were observed with keratinocyte cultures switched to high calcium conditions for only 9 h (not shown). ( B ) Primary keratinocytes derived from fyn −/− mice and wild-type littermates were exposed to high calcium concentrations (2 mM) for 9 h. Cultures were examined as such ( left panels ) or after treatment with dispase for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the fyn −/− cultures already at this time. There were no cells missing in the monolayer of fyn knockout keratinocytes before dispase treatment. As we previously reported ( Calautti et al., 1995 ), the fyn knockout keratinocytes fail to stratify and are larger than normal, which explains the different morphological appearance of these cultures relative to the wild-type controls even before dispase treatment. Similar results were observed in two other independent experiments. Bar, 60 μm.
Figure Legend Snippet: Decreased strength of calcium-induced cell adhesion as a consequence of tyrosine kinase inhibition or lack of the Fyn kinase. ( A ) Primary keratinocytes under growing conditions were either tested as untreated controls ( Ctr ) or pretreated for 2 h with Genistein. Incubation was then continued for additional 24 h under high calcium conditions. Cultures were examined as such ( left panels ) or after dispase treatment for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the tyrosine kinase inhibitor-treated cultures. No such areas were evident in control cultures even after prolonged dispase exposure ( > 30 min). Instead, control cells eventually detached from the dish as a confluent sheet. Similar results were observed with keratinocyte cultures switched to high calcium conditions for only 9 h (not shown). ( B ) Primary keratinocytes derived from fyn −/− mice and wild-type littermates were exposed to high calcium concentrations (2 mM) for 9 h. Cultures were examined as such ( left panels ) or after treatment with dispase for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the fyn −/− cultures already at this time. There were no cells missing in the monolayer of fyn knockout keratinocytes before dispase treatment. As we previously reported ( Calautti et al., 1995 ), the fyn knockout keratinocytes fail to stratify and are larger than normal, which explains the different morphological appearance of these cultures relative to the wild-type controls even before dispase treatment. Similar results were observed in two other independent experiments. Bar, 60 μm.

Techniques Used: Inhibition, Incubation, Derivative Assay, Mouse Assay, Knock-Out

Related Articles

Incubation:

Article Title: Human Keratinocytes Cultured on Collagen Matrix Used as an Experimental Burn Model
Article Snippet: .. These fragments were placed in a tube containing 30 mL of dispase solution (Boehringer Mannheim cat. No. 165859) and incubated overnight at 4°C for 15 hours. ..

Article Title: Tyrosine Phosphorylation and Src Family Kinases Control Keratinocyte Cell-Cell Adhesion
Article Snippet: .. Dispase Treatment of Mouse Primary Keratinocytes Duplicate 60-mm dishes of confluent keratinocyte cultures in either low or high calcium medium were washed twice in PBS and incubated in 2 ml dispase solution in PBS (from Bacillus polymyxa ; > 2.4 units/ml; Boehringer Mannheim Corp. ) at 34°C. .. Cells were analyzed with an inverted transmission microscope at 5-min intervals, for > 35 min. Quantification of this assay was performed as follows: after 35 min of dispase treatment, cells from each sample, including single cells released into suspension and detached sheets of cells, were collected by scraping, and then washed twice in PBS.

Transferring:

Article Title: Focal adhesion kinase knockdown modulates the response of human corneal epithelial cells to topographic cues
Article Snippet: .. Briefly, scleral and limbal areas of the corneas were trimmed and the remaining tissue was immersed in dispase solution (1.2 U ml−1 , Boehringer, Mannheim, Germany) at 37 °C for 4 h. Corneal cells were removed by gentle rubbing of the anterior surface with a pipette tip. ..

Article Title: The scale of substratum topographic features modulates proliferation of corneal epithelial cells and corneal fibroblasts
Article Snippet: .. Briefly, scleral and limbal areas of the corneas were trimmed and immersed in dispase solution (1.2 U/mL, Boehringer, Mannheim, Germany) at 37°C for 4 h. Corneal cells were removed by gentle rubbing of the anterior surface with a pipette tip and the resulting cell suspensions were pooled and collected. .. Cell suspensions were centrifuged and resuspended in Epilife basal medium (Cascade Biologics) with the following growth supplements: bovine serum albumin, bovine transferrin, hydrocortisone, recombinant human insulin-like growth factor type-1, prostaglandin, and recombinant human epidermal growth factor.

Mouse Assay:

Article Title: Regulation of the Spatial Organization of Mesenchymal Connective Tissue
Article Snippet: .. Cultures of confluent keratinocytes (35-mm dishes) were detached as intact epithelia with Dispase II solution (1.2 U/ml for 1 hour at 37°C; Boehringer Mannheim) and grafted to athymic mice as described. .. Briefly, the detached epithelium was placed basal side up on a small piece of Silastic membrane (thickness, 0.005 inches; Dow Corning, Arlington, TN).

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    Boehringer Mannheim dispase solution
    Quantitative measurements of keratinocyte cell adhesion as determined by a novel <t>dispase-based</t> assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.
    Dispase Solution, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 92/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dispase solution/product/Boehringer Mannheim
    Average 92 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    dispase solution - by Bioz Stars, 2021-01
    92/100 stars
      Buy from Supplier

    90
    Boehringer Mannheim collagenase dispase solution
    Quantitative measurements of keratinocyte cell adhesion as determined by a novel <t>dispase-based</t> assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.
    Collagenase Dispase Solution, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 90/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/collagenase dispase solution/product/Boehringer Mannheim
    Average 90 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    collagenase dispase solution - by Bioz Stars, 2021-01
    90/100 stars
      Buy from Supplier

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    Quantitative measurements of keratinocyte cell adhesion as determined by a novel dispase-based assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.

    Journal: The Journal of Cell Biology

    Article Title: Tyrosine Phosphorylation and Src Family Kinases Control Keratinocyte Cell-Cell Adhesion

    doi:

    Figure Lengend Snippet: Quantitative measurements of keratinocyte cell adhesion as determined by a novel dispase-based assay. ( A ) Primary keratinocytes under low calcium conditions ( Low Ca ) or treated with calcium for 2 or 9 h ( Ca ) were incubated with dispase for 35 min as described in Materials and Methods. Data are expressed as percentage of single cells released by mechanical disruption after dispase treatment versus total number of cells recovered after subsequent treatment of the same samples with trypsin. ( B ) A similar assay was performed with keratinocytes under low calcium conditions or incubated with calcium for 9 h ( Ca ) in the absence or presence of Genistein (100 μM) ( Gen ), Thyrphostin (100 μM) ( Trph ), or PP1 (1 μg/ml). Cells were preincubated with the inhibitors for 2 h before calcium treatment. ( C ) Keratinocytes under low calcium conditions were infected with a control adenovirus expressing the green fluorescent protein ( Ad-GFP ) or a virus expressing a constitutively active form of c-src ( Ad-src ). 24 h after infection, cells were switched to high calcium medium and incubation was continued for additional 24 h ( Ca ). Cells were analyzed by the dispase assay as before, together with control uninfected cells kept in either low or high calcium medium for 24 h.

    Article Snippet: Dispase Treatment of Mouse Primary Keratinocytes Duplicate 60-mm dishes of confluent keratinocyte cultures in either low or high calcium medium were washed twice in PBS and incubated in 2 ml dispase solution in PBS (from Bacillus polymyxa ; > 2.4 units/ml; Boehringer Mannheim Corp. ) at 34°C.

    Techniques: Incubation, Infection, Expressing

    Decreased strength of calcium-induced cell adhesion as a consequence of tyrosine kinase inhibition or lack of the Fyn kinase. ( A ) Primary keratinocytes under growing conditions were either tested as untreated controls ( Ctr ) or pretreated for 2 h with Genistein. Incubation was then continued for additional 24 h under high calcium conditions. Cultures were examined as such ( left panels ) or after dispase treatment for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the tyrosine kinase inhibitor-treated cultures. No such areas were evident in control cultures even after prolonged dispase exposure ( > 30 min). Instead, control cells eventually detached from the dish as a confluent sheet. Similar results were observed with keratinocyte cultures switched to high calcium conditions for only 9 h (not shown). ( B ) Primary keratinocytes derived from fyn −/− mice and wild-type littermates were exposed to high calcium concentrations (2 mM) for 9 h. Cultures were examined as such ( left panels ) or after treatment with dispase for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the fyn −/− cultures already at this time. There were no cells missing in the monolayer of fyn knockout keratinocytes before dispase treatment. As we previously reported ( Calautti et al., 1995 ), the fyn knockout keratinocytes fail to stratify and are larger than normal, which explains the different morphological appearance of these cultures relative to the wild-type controls even before dispase treatment. Similar results were observed in two other independent experiments. Bar, 60 μm.

    Journal: The Journal of Cell Biology

    Article Title: Tyrosine Phosphorylation and Src Family Kinases Control Keratinocyte Cell-Cell Adhesion

    doi:

    Figure Lengend Snippet: Decreased strength of calcium-induced cell adhesion as a consequence of tyrosine kinase inhibition or lack of the Fyn kinase. ( A ) Primary keratinocytes under growing conditions were either tested as untreated controls ( Ctr ) or pretreated for 2 h with Genistein. Incubation was then continued for additional 24 h under high calcium conditions. Cultures were examined as such ( left panels ) or after dispase treatment for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the tyrosine kinase inhibitor-treated cultures. No such areas were evident in control cultures even after prolonged dispase exposure ( > 30 min). Instead, control cells eventually detached from the dish as a confluent sheet. Similar results were observed with keratinocyte cultures switched to high calcium conditions for only 9 h (not shown). ( B ) Primary keratinocytes derived from fyn −/− mice and wild-type littermates were exposed to high calcium concentrations (2 mM) for 9 h. Cultures were examined as such ( left panels ) or after treatment with dispase for 5 min ( right panels ). Arrows , the focal areas of cell detachment that occurred in the fyn −/− cultures already at this time. There were no cells missing in the monolayer of fyn knockout keratinocytes before dispase treatment. As we previously reported ( Calautti et al., 1995 ), the fyn knockout keratinocytes fail to stratify and are larger than normal, which explains the different morphological appearance of these cultures relative to the wild-type controls even before dispase treatment. Similar results were observed in two other independent experiments. Bar, 60 μm.

    Article Snippet: Dispase Treatment of Mouse Primary Keratinocytes Duplicate 60-mm dishes of confluent keratinocyte cultures in either low or high calcium medium were washed twice in PBS and incubated in 2 ml dispase solution in PBS (from Bacillus polymyxa ; > 2.4 units/ml; Boehringer Mannheim Corp. ) at 34°C.

    Techniques: Inhibition, Incubation, Derivative Assay, Mouse Assay, Knock-Out