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9

I agree with @Jeremias Brand's answer. Pretty much you will have to forget about fluorescence microscopy... you can probably find some dusty old one on eBay in your price range, but it probably won't be any good. However, the good news is that seen that in your comment you mention a) plants, b) blood, c) liquids such as wine, d) food? transmitted ...


7

With all respect I think the accepted answer underestimates the quality of current inexpensive instruments. What I have found comparing images on my recently acquired $400 scope to those produced by top-end Nikons is that it produces images which are aesthetically less appealing but nearly identical in detail. Mostly I have used it for fungi, which are ...


7

The theoretical resolution for an optical microscope depends on the wavelength used, but is close to 0.22 um (220 nm). Confocal microscopy does not greatly improve axial resolution; it dramatically improves the in-plane resolution (x and y directions, by excluding extraneous light). Theres not a lot to do to "cope" with that using those techniques. If you ...


6

This really depends on the application you have in mind. As with other precision instruments there is a huge range of qualities and applications. If you just want brigth field illumination and look at relatively big things ( approx 100 microns) then you could find something decent for the price you mention if you buy used. But if you want more complex ...


5

For fluorescence immunocytochemistry, there are 3 main incubations that determine duration of the procedure. Generally it takes 2 days to stain your cells before you can visualize them. Once cells or tissue are fixed onto glass, you first incubate with a primary antibody to directly target a specific antigen(s), usually overnight. After primary labelling, a ...


5

You could also have a look at Leica Microsystems' Science Lab. You will find lots of articles and tutorials on the different microscopy methods, ranging from basic microscopy knowledge to specific know-how, including latest information on STED: www.leica-microsystems.com/science-lab For example: gCW-STED Microscopy: When the Arrival Time of a Photon Matters ...


4

You could try SYTO Orange. Unlike SYTOX, which is a fixed-cell marker, SYTO works with both live and fixed cells.


3

A couple of review articles you could read up on. 1) Leung BO, Chou KC. Review of super-resolution fluorescence microscopy for biology. Appl Spectrosc. 2011 Sep;65(9):967-80. 2) Huang B, Bates M, Zhuang X. Super-resolution fluorescence microscopy. Annu Rev Biochem. 2009;78:993-1016.


3

DAPI and Hoechst 33342 (there are different Hoechst dyes, 33342 is one of the most commonly used) have very similar spectral characteristics. The only point is that DAPI is much better excited at 405 nm than Hoechst. Some microscopes and flow cytometers nowadays have 405 nm lasers or LEDs instead of UV sources, so this can become relevant. The main ...


3

The Hoechst 33342 dye is similar to DAPI in that both are UV-excited, minor groove-binding, and emit signals proportional to total DNA content. Both are maximally excited around 355 nm and emit around 460 nm. A UV light source is required, which may harm the cell. However, this is only a risk with Hoechst, as DAPI requires the cells to be fixed and/or ...


3

Extracellular matrix (ECM) fluoresces, especially Collagen and Laminin. The maximum is in the DAPI and FITC channels and the fluorescence becomes weaker towards longer wavelengths. However, since the coat on the TC flasks is very thin, I would not expect this to be a problem. The best thing is just to try it. There is also a quite famous document available ...


2

Very popular microscopes for bio work these days are the inverted style, These allow you to view samples without preparing slides. http://en.wikipedia.org/wiki/Inverted_microscope http://www.biotechequipmentsales.com/equipment-for-sale/details/564/14/microscopes/nikon-eclipse-ts100


2

My main concern here is why you are using suspension-culture selected cells for an adherent cell-based assay? Is it possible to obtain a clone of adherent HT1080? Depending on what you are assaying with your protein of interest, the cell biology may be altered changing from suspension/adherent culture systems. Here are my experiences using PC12 cells in ...


2

MitoSOX looks pretty cool. I've never used it myself, but I'm sure you could see some cool mitochondrial dynamics going on. Its a bit pricey though, but all dyes are going to be. Even at only 8 hours you should probably be able to see a few divisions, which would be cool with hoechst.


1

As far as I can see from different articles, this seems to be a study in progress. Dr Kloock from the California state University states that scorpions might be more active when not exposed to UV light. If not fluorescing, they know that they are hidden and have a lesser chance of being preyed on. He argues that scorpions are less active on full moon nights ...


1

I'm going to try to lay out some basic definitions here in as plain language as I can find. Its difficult to study enzymes when they are outside the cell, where they may behave quite differently in different contexts. We categorize a given enzyme in its class by kcat, Km and by mechanism (the sort of reaction they catalyze. kcat is sort of a maximum ...


1

Well you can always use a piece of white circular paper and draw the outline of the light onto it since it should be visible. This is a quick and dirty method that can be improved by taking a photo of the light with a camera if you have one and then comparing the distance of the edge of the paper measured with a ruler to that of the circle of light.


1

There are only 3 classes of fluorescent proteins proteins known I think. Two of them are relevant to your needs maybe. The first are usually called 'fluorescent proteins'. These proteins are usually derived from the A victoria jellyfish. Its entirely passive in its fluorescence, but has been tuned to many levels of quantum yield and color. There was ...


1

I found this paper. Zhang GJ et al. (2009) In vivo optical imaging of LacZ expression using lacZ transgenic mice. Assay Drug Dev Technol.7:391-9. doi: 10.1089/adt.2009.0195. Abstract beta-Galactosidase (beta-gal) (encoded by the lacZ gene) has been widely used as a transgene reporter enzyme. The ability to image lacZ expression in living ...


1

Yes, they will both affect pH. In addition to the above comment, both, but methanol especially dehydrates samples - removal of water will most definitely affect the pH in organelles. This will be time dependent. Both duration of fixation, and time after fixation before you assess the cells will affect the pH of the organelles. Because you fix the cells in ...


1

In my opinion, cell fixation shouldn't change the pH. However unbuffered formalin will oxidize and lower the pH, but using PBS should buffer around pH 7. Maybe Glutaraldehyde fixation would also be an option, if the others are not working... I found this website by leica very useful, maybe it will also help you: ...


1

Have you looked into trying the DropArray microplates? They're a new technology where you don't need to centrifuge your cells to wash them...an old colleague of mine works for the company and they're pretty cool about demoing the products/instruments for you. It's http://www.curiox.com/. I think most people that use the system are using suspension cells- ...



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