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fysio lab's Content

There have been 11 items by fysio lab (Search limited from 19-January 19)

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#128676 Culturing cells on a slide

Posted by fysio lab on 08 February 2012 - 07:42 AM in Cell Biology

Most adherent cells should grow on glass just fine with no poly L lysine treatment. Some tricky cells (e.g. 293) might require poly L lysine so that they don't spontaneously detach. Chamber slides are nice, since they will allow you to use less reagents for later processing steps. In the old days, we used to just drop sterile cover slips into 6-well plates, and cells would attach just fine to the cover slips. If doing confocal microscopy, the thickness of the coverslips may be crucial, so check with your local confocal expert prior to choosing cover slips. If you are trying to visualize a fluorescent protein expressed by your cells, you won't need to fix the cells. If you are going to be doing staining with antibodies, you will need to do a fixation/permeabilization step. I personally like to use one that involves 3.7% formaldehyde (in PBS) at 37C for 10 minutes followed by 100% of -20C Methanol for 30 minutes.

Chamber slides are indeed very handy...but you need to know which type of CSLM you have first. Is it an upright or an inverted?

#123869 Help with Ethidium Bromide Contamination

Posted by fysio lab on 18 November 2011 - 04:53 AM in Biochemistry

We once checked a DNA-electrophorese-room for contamination of etbr with a strong UV-lamp. The view was dramatic! Walls and wastebins were glooming.
Maybe you can check with a UV-lamp for severity of contamination?
Good luck

#118760 Imaging GFP in S. pombe

Posted by fysio lab on 02 September 2011 - 12:03 AM in Cell Biology

you said you're loading your cells on a coverslip. Do you coat this and is it possible that this is giving background? We pipet our yeasts on an microscopyglass and put a coverslip on top (not ideal cause our yeasts tend to 'swim). But keep in mind that coating solutions can give background
Good luck

#117778 How do you coat coverslips before seeding cells?

Posted by fysio lab on 19 August 2011 - 01:25 AM in Tissue and Cell Culture

we put non-sterile coverslips in a glass petridish, wrap that in alu-foil, autoclave it and before use we put the CS in the wells with sterile forceps. After that we put our coating solution on the CS (in our case 2% gelatin), we leave it 30' in the incubator. Then we remove the gelatin and let it dry for a few minutes in the flow, lid open... meanwhile we split and count our cells. All together this doesn't take much time.
Good luck

#117197 Marker pens for glassware

Posted by fysio lab on 12 August 2011 - 01:32 AM in Lab Equipments

we were looking for the same kind of pens a while ago. We found good ones from a brand called Edding (751 paint marker, red). Hope this helps?

#115029 gel question

Posted by fysio lab on 14 July 2011 - 01:04 AM in Molecular Biology

Phage is right. Next time you better cut out your band and store the slice dry(!) in the freezer...then you can keep it as long as you want. This one you better proceed with.
Good luck

#113430 localization

Posted by fysio lab on 24 June 2011 - 12:38 AM in Cell Biology

Looks to mitochondria to me...stressed mitochondria curl up: so how did you treat your cells? fixed, non fixed? You can use Mitotracker, it's a good dye.
Good luck

#113131 Opening a sterile reagent out of the tissue culture hood

Posted by fysio lab on 21 June 2011 - 12:54 AM in Tissue and Cell Culture

If you really want to use it: fill a tube with LB and a tube with YP, add your dmso and incubate at the appropiate temperatures, check after a few days for growth

#100785 need a math check

Posted by fysio lab on 16 February 2011 - 01:26 AM in General Lab Techniques

your way is also correct, different people--> different way of calculations ;)

#100784 mycoplasma detection kit

Posted by fysio lab on 16 February 2011 - 12:32 AM in Cell Biology

we use plasmotest from invivogen...works with 'reportercells' no pcr! reduces the chance of falsepos or falseneg
Good luck!

#100644 need a math check

Posted by fysio lab on 15 February 2011 - 01:35 AM in General Lab Techniques

i think: if you have 0.1 ng/ul at 10 ul--> is 1 ng. You add 5 ul of 0.5 ng/ul, so you add 2.5 ng. Your total is 3.5 ng at 15 ul, so 3.5/15= 0.233333 ng/ul

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