9
votes
Alternative dyes for Gram staining
As you may have realized, crystal violet can be replaced by a lot of dyes since the ethanol will wash out the dye from the gram negative cells. Methylene blue is a nice choice - malachite green may ...
- 201
9
votes
Accepted
"X" in stock solutions
1x is the final working concentration of the solution (it could be anything depending on the type of the solution). Stock solutions are made at a higher concentration; if it is 10 times more ...
- 35.2k
8
votes
Accepted
Reason for the source of fetal bovine serum
The reason fetal bovine serum (FBS) is used is because it supports a wide variety of cells and really supports their growth. It has a high concentration of growth factors, but also a low ...
- 1,152
6
votes
Accepted
1% water solution of deoxycholic acid. How is it prepared?
According to the sigma document you referenced (emphasis added by me):
"The free acid is soluble at 0.24 g/L in water at 15ºC, while the sodium salt is soluble at >333 g/L in water at 15ºC. ...
- 4,571
6
votes
Accepted
Is there a difference between Luria Broth and Lysogeny Broth?
Although both names are quite common, both are wrong. In the original paper from 1951 Bertani was studying the lysogeny in E.coli, hence he called his media for this purpose "Lysogeny Broth" or LB in ...
- 51.2k
5
votes
Accepted
Why do many DNA solutions contain additional compounds?
DNA in pure water.
The only time that nucleic acids would encounter pure water would be in a laboratory setting--for example after an oligonucleotide is synthesized in vitro, the protecting groups are ...
- 3,517
5
votes
Is NADPH salt still usable after an hour at room temperature?
In my experience, half an hour at room temperature will make absolutely no difference. Boehringer Mannheim (now part of Roche), who at one time supplied the best NADPH, used to recommend storage at ...
- 4,668
4
votes
Is NADPH salt still usable after an hour at room temperature?
You could measure OD at 340nm. If OD340 is much lower than expected, NADPH is oxidized and does not have much biochemical activity.
http://www.bmglabtech.com/media/35216/1043734.pdf
- 1,116
4
votes
Is there a reliable source for storage and stability of reducing agents like DTT?
Thiol-Based Reducing Agents
The instability of thiol-based reducing agents in solution is due to their propensity to form disulfide bonds by the following half reaction:
$$\ce{2RSH -> RSSR + 2H+ +...
- 17.6k
4
votes
DNA quantification in a high school bio lab
Within some linear range, the intensity of stained DNA bands in a gel is directly proportional to their mass. By making a standard curve from bands of known mass, the mass of unknown bands can be ...
- 17.6k
4
votes
How pricey does the media need to be for a typical bacterial culture experiment?
It looks like Merck have different levels of quality control for their products. MQ 100 being the lowest up to MQ 600 being the highest. From what I can see the MQ200 product here has defined ...
- 431
3
votes
How standardized is lab-grade skim milk?
There is a degree of variability in milk, and in some cases this cause problems in protein work. Despite being a standard for ELISAs, it's not known to be the most reliable blocking buffer. As far as ...
- 191
3
votes
Accepted
Using autoclaved store-bought distilled water for labwork?
Yes, distilled water would be absolutely fine for your needs. You really don't need ultrapure water except for the most sensitive of applications. You also don't need to autoclave the distilled water ...
- 17.6k
3
votes
How pricey does the media need to be for a typical bacterial culture experiment?
The answer depends on the application and level of oversight/compliance you require. Most research labs I've seen use de-ionized or even reverse-osmosis water (from a tap) for most culture media ...
- 4,571
3
votes
Accepted
Ordering Primers: When to choose wet vs dry delivery format?
Pros of wet:
No need to rehydrate to use.
For e.g. 96-well plates, this can be a big deal as it's a pain to do this for large numbers of samples.
More accurate concentrations (oligo manufactureres ...
- 8,738
3
votes
Accepted
Why is it advised to avoid bubble formation during mixing?
A "bubble" per se would do no harm unless you have cells (without walls like mammalian cells) in the suspension. If you agitate the protein mixture vigourously then it may lead to denaturation of ...
- 35.2k
3
votes
Agarose gel Ladder smear
Are you sure that your TBE is at the right dilution? And that you're using TBE to make the gel and also as the buffer for running? That sort of wavy line looks like what you might get if you'd made ...
- 41
3
votes
Is NADPH salt still usable after an hour at room temperature?
I think when they sent it to you without dry ice, it is probably OK to store it at room temperature.
Sigma seems to advise on their NADPH to store dehydrated NADPH at room temperature, while they ...
- 52.2k
2
votes
Trypsin and cell culture
It would be stressful for cells to be trypsinized 24 hours after seeding. After cells are plated, there is a lag time to start growing. Perhaps, during the lag time, physiological state of cells is ...
- 1,116
2
votes
Agarose gel Ladder smear
Try going even higher with the agarose concentration, as it looks like you have decent separation of the small bands and poor separation of the large bands. Also try a fresh batch of agarose.
I ...
- 677
2
votes
Is there a protocol for freezing and thawing Bacillus subtilis cells?
While I have never done bacterial stock cultures from spores, I don't think it is necessary, as the standard procedure for making bacterial stocks should work without problems.
For that you grow ...
- 51.2k
1
vote
Accepted
Washing buffers for protein identification?
All the unoccupied sites on the membrane are saturated in the blocking step after the transfer of the protein gel to the membrane. You can prepare the antibody solution in pure buffer, but you should ...
- 51.2k
1
vote
How pricey does the media need to be for a typical bacterial culture experiment?
This answer only concerns Streptococcus pneumoniae.
The lab I worked in used cheap ingredients, the same as everyone else claimed to in this field.
The only shocking thing I found was that the ...
- 2,838
1
vote
Use lab agar for cooking
Technically agar has first been used (and still is) in the kitchen and then adopted to the lab.
In the 19th century, gelatin was used in the microbial laboratory. The problem is that gelatin is much ...
- 51.2k
1
vote
FPLC based separation of serum proteins
Firstly, may I suggest 2D electrophoresis as a technique for identifying differences in protein/marker expression levels. This technique will give you greater resolution and ease of comparing your ...
- 122
1
vote
Trypsin and cell culture
Might be considered "not answering the question" but:
Option 1:
Depending on the type of cells you are using, you can use milder methods to detach cells. Example include Versene which is essentially ...
- 121
1
vote
Trypsin and cell culture
It is not advisable to trypsinize the cells too often when you have to maintain them. However, in your case the situation is different.
Time required to attach depends on cell type. In any case ...
- 35.2k
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