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LacZ staining of cells: Difference between revisions
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The bacterial enzyme β-galactosidase (gene LacZ) is frequently used as reporter gene. It can be easily located with a LacZ stain using the artificial substrate X-gal. | |||
This protocol can be used for staining cultured cells containing active LacZ genes. | |||
== material == | == material == | ||
* X- | * X-gal stock is 200 mg/ml in dimethylformamide (DMF) | ||
* K ferri-cyanide ( 50mM )= | * K ferri-cyanide ( 50mM ) =Fe3- is 0,33 g / 20 ml PBS ( Sigma Cat.# P-8131) | ||
* K ferro-cyanide ( 50mM )= | * K ferro-cyanide ( 50mM ) =Fe4- is 0,42 g / 20 ml PBS ( Sigma Cat.# P-9387) | ||
=== staining solution === | === staining solution === | ||
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| MgCl<sub>2</sub> (1M)||20 µl||30 µl||60 µl||100 µl | | MgCl<sub>2</sub> (1M)||20 µl||30 µl||60 µl||100 µl | ||
|- | |- | ||
| K ferri-cyanide ( 50mM ) | | K ferri-cyanide ( 50mM )||1 ml||1,5 ml||3 ml||5 ml | ||
|- | |- | ||
| K ferro-cyanide ( 50mM ) | | K ferro-cyanide ( 50mM )||1 ml||1,5 ml||3 ml||5 ml | ||
|- | |- | ||
| PBS ( up to...)||7,9 ml||8,5 ml||23,5 ml||39,6 ml | | PBS ( up to...)||7,9 ml||8,5 ml||23,5 ml||39,6 ml | ||
