Questions tagged [lab-techniques]

Questions relating to protocols, procedures, and good practice when using laboratory equipment.

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49 views

Why is plasma glucose concentration not double that of whole blood?

It is known that the concentration of plasma glucose is 12% higher than that of whole blood. But since 45-50% of whole blood is red blood cells, shouldn't the plasma glucose be almost double — since ...
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35 views

Control for Bisulfite sequencing

I am wondering how can I control my converted DNA! I converted my DNA with bisulfite then I amplified the converted DNA with specific primers of BSP and purified my product and finally sequenced the ...
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18 views

Minimum number of cycles for effective qPCR quantification?

I have been having an apparent problem with gDNA contamination in my no-reverse transcriptase (NRT) controls by the appearance of fluorescence peaks in my qPCR data. I have/am trying multiple DNase ...
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1answer
900 views

Why shouldn't I clean microscope slides with a paper towel?

I've recently bought a microscope and been looking at a lot of bacteria and such, and have quickly run out of clean slides. I've seen videos and articles saying how you shouldn't clean microscope ...
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1answer
184 views

How do you keep track of past/new lab protocols?

Does there exist a list grouped my subject (e.g. 'microscopy') for keeping track of what protocols have been designed and used? Closest I could find is http://www.protocol-online.org/ but it's fairly ...
2
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1answer
28 views

What do the column labels in this table mean?

This table shows the effect of sound (song and tones) on female birds. However, I'm not sure what the labels (F1 22, P and η2) mean. I've seen the labels on other tables too. (from https://www.ncbi....
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87 views

Would I expect salt precipitate on fibres of DNA in a NaCl water solution?

I just recently conducted an experiment in my biochemistry class where we had to add DNA to distilled water, isotonic saline and 2.5M NaCl. I noticed different physical aspects depending on the ...
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1answer
478 views

How is the gradient in density gradient centrifugation made?

From the literature I read, I came to understand the following. Please correct me if wrong. In CsCl gradient centrifugation, the gradient is achieved by centrifuging the CsCl solution at high rpm, ...
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2answers
27 views

Do PC-12 cells have to be stored in nitrogen?

My lab has been storing their PC 12 cells in the -80 freezer... upon arriving here and investigating a new project I see that it specifically stated to be stored in nitrogen vapor? Is this the ...
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186 views

What are the optimal conditions for storing frozen glycerol stocks of bacteria?

The common method of storing bacteria long-term is to collect saturated liquid culture, such as LB, and mix it with sterile glycerol so as to obtain 10-30% final glycerol concentration. The resulting ...
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1answer
61 views

How to choose right volume of DNA and SYBR Green

I'm testing fluorescence level of a sample having dsDNA, SYBR Green and milli q water but I'm facing difficulty in choosing right volume of dsDNA and cons level of SYBR Green. Details of sample: ...
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53 views

What are these colonies on my Bacillus pumilus lawn?

I am growing Bacillus pumilus and I have plated some on agar plates. Now I can see some colonies on top of the lawn. Does anybody know what they could be? A contaminant? Bacillus pumilus? I have ...
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3answers
249 views

Using nanodrop for analysing biological samples other than nucleotides

I am a 3rd timer postdoctoral fellow with some experience in molecular biology and biochemistry, but major skills in Zoology and Natural History. I am studying some natural extracts, and isolating ...
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163 views

At what g force do bacteria start to pellet down a tube?

I am working with C. elegans and bacteria and I want to get rid of the bacteria they eat by centrifuging the worms without centrifuging the bacteria. I am using a g-force of 600 and the bacteria ...
4
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1answer
173 views

Test of antibiotic A for sensitivity as surrogate of antibiotic B

I am reading the WHO Global Antimicrobial Resistance Surveillance System (GLASS) - Manual for Early Implementation. it mentioned some of the drug-bug combination that will be put under surveillance in ...
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1answer
232 views

Should I use RNAseZAP on my bench and pipettes before RNA extraction?

Should I spray RNAseZAP on my bench and pipettes before working with RNA? I am regularly using RNase at my bench and would like to work with RNA as well. Could traces of RNase on my equipment degrade ...
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27 views

How to convert a test tube assay to microplate assay?

I am trying to measure Ascorbic acid levels in tissue using Omaye et al protocol (Methods in Enzymology, Volume 62, 1979, Pages 3-11). This method is based on reduction of ferric chloride and ...
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36 views

What is known and what can be measured of biofilms?

Context for the question I am a mathematics researcher working on the mathematics of imaging and parameter estimation (inverse problems). I found an interesting diffusion equation that models the ...
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1answer
136 views

DNA quantification in a high school bio lab

I'm working on a project in a high school bio lab (so limited resources), and I need a way to quantify the concentration of DNA in a PCR product. I can't use spectrophotometry (cheap ...
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1answer
39 views

Why marker and control are loaded in the beginning or ending lane of the gel?

I have seen gel images in several literature. Almost in all, markers and controls are loaded in the extreme lanes (before 1st sample or after last sample) of the gel. I am just curious, is there any ...
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2answers
2k views

Is working with (nitrile) gloves around a bunsen burner safe?

Recently I have started in a new microbiology lab, and with a new lab come new habits. When I was working with my bacterial (liquid) culture next to the flame (Bunsen burner) wearing nitrile gloves, ...
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44 views

Can anyone expolain this? Cell culture vials have strange hair-like condensation patterns

What is the hair-like polymer that condenses on the caps of the plastic cell vial? Inside vial are cells in a mixture of FBS and DMSO. The vials were frozen slowly in a special isopropanol-filled ...
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1answer
32 views

HELP hepes preparation [closed]

I need help: I would like to prepare an HEPES solution, without using the powder. It is possible? I remember that I did it once, but maybe I'm wrong...does someone know a protocol for HEPES ...
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1answer
147 views

Optimization of Signal Peptides

We are looking at modifying the signal peptide of a receptor in a common immortalized human cell line. The cell line already expresses an unusually high amount of the protein, but much of it is not ...
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0answers
192 views

How to keep Paraffin Sections from falling apart?

I've been trying to make 5 uM sections of injured mouse spinal cords and stain the tissue with Luxol Fast Blue and H&E. At first I was removing the bones from around the spinal cord, but the ...
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2answers
930 views

Agarose gel Ladder smear

We are having trouble with agarose gel electrophoresis. It used to work a couple of months ago but now the ladder always look smeared. We switched the components (1x TBE, 100bp ladder, different type ...
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84 views

What happens during fixation of Nucleic acids on nylon membrane

In Southern blotting technique, fixation of DNA to nylon membrane is a step prior to hybridisation with probe. It is done by baking at 80ºC and according to this cross linking between DNA and nylon ...
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1answer
3k views

What is the role of glucose in plasmid isolation?

My understanding is that glucose is used in the resuspension solution to prevent cells from bursting by maintaining the appropriate osmotic pressure. Why do we even bother doing this? The cells are ...
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1answer
214 views

How can the melting temperature of PCR primers be so far below the extension temperature?

Since our primer TM is lower than the extension temperature by design, (and often by a significant amount) shouldn't the primers be released from the template at extension temperatures? Now, I have ...
3
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2answers
1k views

Why does high EDTA concentration cause swelling of platelets?

According to this textbook, high ( >2mg/ml of blood) EDTA concentration causes swelling of platelets which lead to their fragmentation. I know that high concentration of EDTA increases plasma ...
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0answers
37 views

How urination prior to blood test may give error?

Source: Textbook of Practical Physiology, 4th Edition according to it urination within 30 minutes is a precollection factor that may alter the results of blood test. I am unable to comprehend how ...
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1answer
79 views

How to watch a Zebrafish embryo in detail?

How much microscope zoom would I need to watch the development of a Zebrafish embryo in certain detail? Thanks guys!
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13 views

Propagating errors for percentage reduction

Need a bit of help for this particular problem. I have three averages collected from three genotypes (A, B, and C) and their associated SD (a, b, and c). A is my control. I have been asked to ...
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35 views

Nucleic acid extractions

Is it fine to leave pulverized plant tissue in CTAB buffer at -80 degrees over night? will the buffer remain effective to generate good quality nucleic acids?
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101 views

What are preparation mistakes that could make red blood cells appear crenate in isotonic solution?

Thank you for your time. I separated red blood cells from a whole blood sample purchased from a blood bank. In isotonic solution, the red cells appear crenate. I would like to know if there is ...
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1answer
173 views

Help with preparing Congo Red TSA

So I want to make Congo Red TSA. My instructor gave some hints but I still have some questions: I start by making two stock solutions: 20 mg/ml of congo red and coomassie in two different falcon ...
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1answer
131 views

Is it possible to produce a tangible mass of DNA/RNA using a PCR reaction?

A Polymerase Chain Reaction (PCR) is a method of amplification which enables one to produce many copies of a certain DNA/RNA strand for many applications (as described in the link above). I have ...
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78 views

Plasmid transformation yields only empty vectors

I have to electroporate pMAD cloned with an insert of 7.4 kb in my S. aureus isolate. When I electroporated in RN4220 I had no problems but when I put it in my isolate I just recover few colonies and ...
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55 views

plausible science for why an enzyme breaks down a substrate better at 4 degrees rather than 20 degrees [closed]

An enzyme was found to have an optimal temperature of 20 degrees. The same enzyme in sheep liver extract was found to have an optimal temperature of 4 degrees. What are some of the factors that can ...
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2answers
1k views

Lab marker for labelling cell culture plastics?

I am a lab manager trying to ask other lab biologists what brand/make of markers their labs use for labeling tissue culture plastics, which need to be repeatedly wiped down with 70% ethanol to ...
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1answer
779 views

Are vacuum centrifuges the same as normal centrifuges?

I came across a method that used a vacuum centrifuge: Carvalho et al. 2012. The diversity, antimicrobial and anticancer activity of endophytic fungi associated with the medicinal plant ...
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1answer
4k views

How to convert skin temperature to core temperature?

As opposed to regular invasive thermometers, there are non-invasive IR thermometers to measure the temperature. For example Thermofocus. I read in many sources that these IR thermometers only measure ...
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1answer
340 views

Why is thrombin time (TT) normal range longer than prothrombin time (PT)?

Reference range for the TT is longer than that of the PT.
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1answer
107 views

correction of read counts for spikes

I have 4 RNA-seq samples (1st is control and the other 3 are treatments) and 5 spikes. I used different concentrations for 5 spikes but over different samples, I used similar concentration. for ...
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1answer
685 views

Problems with SDS-PAGE, no separation - what could be the problem?

I work in a lab were we have a common stock of 30% Acrylamide, TEMED, TRIS-HCl (pH6.8 and pH 8.8.) and a 10% (w/v) SDS solution pH 7.2. We use the recipe for loading buffer (which we of course mix ...
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212 views

Why are my assay absorbance values very low?

I am culturing SH-SY5Y cells in 96-well plates at a seeding density of $1.2 \times 10^5$ cells/mL. I differentiate the cells with retinoic acid for 6 days before giving them treatments. The cells look ...
3
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1answer
163 views

FPLC based separation of serum proteins

I am working on a project which involves bio-marker discovery in neuro-developmental disorders. I have an akta-prime FPLC instrument in my lab but I do not know how to use it. According to theory, ...
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1answer
881 views

Aseptic technique: a single technique, or any technique fulfilling criteria, or collection of said techniques?

My friend just used the following sentence in her lab report: Aseptic technique is used in this step: I immediately felt something was wrong. She used the term as if it were a specific technique. ...
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1answer
146 views

How do I improve expression using the GAL1 promoter in S. cerevisiae above the “leaky” level I'm currently seeing?

I am attempting to express GFP in S. cerevisiae using the GAL1 promoter. I always grow an uninduced (in dextrose based SD medium) culture alongside my induced (in galactose based SD medium) culture. ...
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1answer
249 views

During the process of plasmid isolation, how can we be sure that only plasmid DNA has been isolated and there is no chromosomal DNA?

We separate the membrane and all other cell components, but how is it that only plasmid DNA is isolated and not chromosomal DNA?