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[1.] Iam/Fragment 026 01 - Diskussion
Zuletzt bearbeitet: 2014-03-12 20:16:24 Graf Isolan
Fragment, Gesichtet, Iam, KomplettPlagiat, SMWFragment, Schutzlevel sysop, Sheikh 2006

Typus
KomplettPlagiat
Bearbeiter
Graf Isolan, Singulus
Gesichtet
Yes.png
Untersuchte Arbeit:
Seite: 26, Zeilen: 1ff (complete)
Quelle: Sheikh 2006
Seite(n): 25-26, Zeilen: 25:15-25 - 26:1-25
3.1.2.b. Preparation of the hepatocyte suspension

After perfusion, the liver was excised and transferred into a sterile glass beaker filled with culture medium M 199 with additives. Glisson’s capsule, i. e. collagen tissue around the liver, was carefully removed and discarded. To obtain a cell suspension, the tissue was disrupted mechanically using sterile forceps. Connective tissue and remainder of the liver capsule as well as big cell aggregates were removed by filtration of the primary cell suspension through a nylon mesh (pore-size 79 μm). Non-parenchymal cells and cell debris were removed by numerous selective sedimentations (20 g, 2 min, 4°C) in wash medium. After the last centrifugation, hepatocytes were suspended in medium M 199 with additives. 50 ml of M 199 was added per 1 g of wet weight of the sedimented cells; the cell suspension typically had a density of about 106/2.5 ml.

3.1.2.c. Media and solutions for hepatocyte preparation and culture

All media and solutions for cell culture were prepared in double distilled water, further purified by sterile filtration and stored at 4°C. All solutions were prepared not more than one day before the isolation.

Krebs-Ringer stock solution

    Final concentration
NaCl   120 mM
KCl   4.8 mM
MgSO4×7H2O   1.2 mM
KH2PO4   1.2 mM
NaHCO3   24.4 mM

The solution was equilibrated with carbogen and pH was adjusted to 7.35 [sic]

Pre-perfusion medium (prepared in 1X Krebs-Ringer solution)

    Final concentration
EGTA   0.25 mM

Collagenase perfusion medium (prepared in 1X Krebs-Ringer solution)

    Final concentration
HEPES   15 mM
CaCl2×2H2O   4 mM
[Collagenase 50 mg

The medium was prepared directly prior to isolation, equilibrated with carbogen for 30 min and finally sterile filtered.]

[Page 25]

3.1.1.II Preparation of the hepatocyte suspension

After perfusion, the liver was excised and transferred into a sterile glass beaker filled with culture medium M 199 with additives. Glisson’s capsule, i.e. collagen tissue around the liver, was carefully removed and discarded. To obtain a cell suspension, the tissue was disrupted mechanically using sterile forceps. Connective tissue and remainder of the liver capsule as well as big cell aggregates were removed by filtration of the primary cell suspension through a nylon mesh (pore-size 79 μm). Non-parenchymal cells and cell debris were removed by numerous selective sedimentations (20 g, 2 min, and 4°C) in wash medium. After the last centrifugation, hepatocytes were suspended in medium M 199 with additives. 50 ml of M 199 was added per 1 g of wet weight of the sedimented cells; the cell suspension typically had a density of about 106/2.5 ml.

[Page 26]

3.1.1.III Media and solutions for hepatocyte preparation and culture

All media and solutions for cell culture were prepared in double distilled water, further purified by sterile filtration and stored at 4°C. All solutions were prepared not more than one day before the isolation.

Krebs-Ringer stock solution
  For 1l Final concentration
NaCl 7 g 120 mM
KCl 0.36 g 4.8 mM
MgSO4×7H2O 0.296 g 1.2 mM
KH2PO4 0.163 g 1.2 mM
NaHCO3 2.016 g 24.4 mM
dd H2O to 1 l  
The solution was equilibrated with carbogen and pH was adjusted to 7.35 [sic]
Pre-perfusion medium
  For 1 l Final concentration
EGTA 95.1 mg 0.25 mM
Krebs-Ringer stock solution to 1 l  
Collagenase perfusion medium
  For 100 ml Final concentration
HEPES 360 mg 15 mM
CaCl2×2H2O 58.8 mg 4 mM
Collagenase 50 mg
Krebs-Ringer stock solution to 100 ml  
The medium was prepared directly prior to isolation, equilibrated with carbogen for 30 min and finally sterile filtered.
Anmerkungen

Though identical nothing has been marked as a citation.

Typical C&P-mistake: The author has overlooked that the "6" in "106/2.5 ml" is an exponent and not just another digit.

Sichter
(Graf Isolan) Schumann


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