ResearchGate Q&A lets scientists and researchers exchange questions and answers relating to their research expertise, including areas such as techniques and methodologies.
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- How do I get whole cell patch clamp in cells deep within the slice?
A small, scattered sub-population of cells in the slice are labeled, and I'm trying to record from them using WC patch clamp. The problem is most of them are deep within the slice, and while I can identify them by DIC, I can hardly see their exact outline or the pipette tip (when going this deep). I think that's why so far I didn't manage to get any stable WC configuration from these cells. I have no problem with patching random superficial cells. Does anybody know any tricks for recording from visually identified cells deep within a slice?
^I've had good luck with the 'tunneling' technique Milan mentions. All I would add is that I've had better luck when I first tunnel with a cleaning pipette pulled a bit thinner than my patch pipette to make a space to drive through and to clean the surface of the cell.Following
- Are journal impact factors updated annually?
A half of my articles (from my beginner years), though in Elsevier and Springer journals, don't have any IF. I regard some of them highly, for the frequent rejections and rigorous revisions they have put me though. Also, with their high downloaded numbers, I think they deserve IF.
Impact factors are updated at least annually. New journals will often track for an impact factor for a few years before being assigned one. Have a look at the website linked below where you can find out more about journal metrics. If you have good data and are confident in your paper, then it is always best to submit to a high impact factor journal that is well esteemed in your field (at least in the first instance).
- What are the major compound-using phenyls that can be separated via stationary phase HPLC?
Phenyl is not widely used in reversed phase HPLC separation as C8 and C18 stationary phases. However it is used in certain cases (compounds), where C8 and C18 do not work.
I've had some success with "difficult" antibiotics (Massimo mentioned the tetracyclines) and vitamins on phenyl columns. Even if your compound isn't "phenolic" in the broader sense, it can help your separation by offering a different selectivity (i.e. shifting your peaks around...)Following
- Why is a simple appraoch to increasing ones own stem cells naturally not taken up more seriously now that we know their importance
Nutrition is still not on the agenda of health care as it has too many uncertainitys. staying with Stem tech simple but scientifically proven with blind study trials is well worth looking at.Following
- Please suggest suitable laser diode to convert up to 20 volt input signal into optical signal?
I have to make a transmitter which will convert electrical signal to optical signal and transmit over fiber up to 5 meter distance the range of input signal will be 10 mvolt to 20 Volt. pls suggest suitable laser diode for this range.
The voltage response of diode lasers is highly non-linear. Converting the input voltage to current, as suggested by David, will greatly improve linearity (and reduce the likelihood of destroying the laser). However, although the optical intensity increases more linearly with current than voltage, laser diodes typically exhibit a lasing threshold current below which the output is very weak. This can be offset in the driver, but the threshold is sensitive to laser junction temperature, so with a constant current drive the output will drift with time as the ambient and laser temperatures change.
David's advice to use a voltage to frequency converter is sound.
If an output with reproducible optical intensity is essential, then some kind of feedback control is highly desirable. Many lasers are provided with a monitor photodiode which delivers a current proportional to the optical output from the laser cavity. Alternatively, a fibre tap coupler and a separate photodiode offers the same functionality. Many commercial laser driver modules support these configurations.
What type of fibre are you using? What is the core size, numerical aperture and operating wavelength? Is it single mode or multi-mode? A collimated laser output may not be the best option. Additional optical components could be needed to couple power efficiently into the fibre core, unless the core is comparable in diameter or larger than the laser beam width. Depending on the type of fibre, laser diodes should be available with matching or compatible pigtail fibres, and this may be a simpler option, especially if you can use a splice or fibre connector.Following
- Why is a change of the pipette resistance normal , when I give a positive pressure?
Hi, I used to work with Cs-based internal solution(Cs-IS). Now, I change to K-Glu based internal solution (K-IS) to record action potential. When I use the K-IS, I found the pipette resistance can change from 3.5M to 4.3M after a positive pressure, but not Cs-IS. When release the positive pressure, the pipette resistance change back to 3.5. I checked the pipette there is no air bubbles or dirty things clog the pipette.
I also checked the osmolarity of K-IS, it is about 285. It should be OK.
I am not familiar with K-IS. Is this normal?
Although debris can cause an increase in resistance as mentioned before, it doesn't sound like that's what you're looking at. To elaborate a bit on the also mentioned ohmic effect (feel free to correct me where necessary):
Recall your VClamp measures voltage and drives a current until the measured current matches the command voltage, resistance is calculated depending on how much current is required to hold a given voltage. Also recall that what is measured and output by the amp is electric current, but what actually flows across the pipette tip are ions, and the current given by the amp is redoxing Ag/AgCl to spit out or chew up Cl- ions making the electric field that pushes ions at the tip.
The junction potential is the result of differing mobilities of ions, depending on the direction of flow (if your extracellular solution was a lower osmolarity, it would flow into your pipette and be the source of the jp, I assume your IS is the lower osm though). Mobility is the result of an ions ability to move in response to an electric field, and since your gluconate ions are much larger than your MeSO4 it is much less mobile. This is essentially electrochemical diffusion, and since the ratio of mobility between cations/anions favors cations more than your Cs-IS. However, pressure adds a current of fluid, and since it is very doubtful the flow out of your pipette tip is laminar, the current and its turbulence have two effects: 1)the current flow of your ions *effectively* magnifies the electric field (imperfectly analogous to an increase in permittivity) by improving Cl- diffusion from the electrode through the pipette and 2)the turbulence 'stirs' the ions - decreasing the impact of electrical mobility and thus effectively decreasing JP.
Then to answer your question, the reason your resistance decreases with positive pressure in your K-IS but not your Cs-IS is that in the case of K-IS, the magnitude of decrease in current required from the amp in the presence of pressure is just much larger because of the difference in mobilities which is then reflected as a larger increase in resistance, whereas with your Cs-IS the decrease in current (and thus increase in resistance) is likely there unless your mobilities are perfectly balanced, just very small due to the mobility of MeSO4.
This is of course a simplified picture, as diffusion is a function of all particles interacting simultaneously, but it works :).
And as said before, yes you have to correct for your junction potential, either in your VClamp protocols or afterwards - usually just a subtraction problem, but you just want to make sure that if you wanted to test a specific voltage in your protocols that they reflect that.
As said above, if i'm way off base i'd love to hear corrections, this is just how I understand it, good luck :)Following
- Why is my PCR cloning technique failing?
I have been trying to clone a 1600 bp GOI into a pET28a vector. My procedure is as follows: I made 6 reactions for 3 different temperatures and 2 GOIs(same gene different code). 1. add 2 microliters of amplified and PCR cleanup GOI. 2. Add .8 microliters pET28a vector. 3. Add 5 microliters of Q5 Polymerase. 4. Add 1.2 microliters ddH2O. 5. Add 1 microliter of engineered reverse primer. 6. The PCR times at temps are as follows: Initialization step-98C for 2 minutes, denaturation at 98C for 30 secs, Annealing- 55,60,65C gradient for 30 seconds, Extension step at 72C for 7 minutes.
I then ran the products of this reaction on a 1% agarose gel using 2 microliters loading dye and 8 microliters of PCR product. I ran a pet28a plasmid in the farthest right lane. Is it possible the products are too concentrated and thats why the DNA isn't leaving the wells?
Our lab has had some success using a new method of PCR based insertions into vectors without the use of restriction enzymes. Theoretically it takes out some of the luck involved with traditional digestion/cloning techniques. New England Biolabs has a protocol and there are some publications online that refer to this method. Although other projects have had success, my GOI has not been as easy.Following
- How can I design a classification network for 6 input parameters and 2 output parameters?
I have 6 parameters each a row matrix of dimension 1 x 5, now i want to consider these as inputs and define 2 outputs. For each parameter i want to set a cut-off and give the appropriate rules to get classified into either of the two outputs. After feeding in the neural network and training it, how to given a new unknown input (i.e. all 6 parameters) and it should be classified into either of the one category. How to design and do this ? My problem case is somewhat similar to the breast-cancer example in the matlab neural network toolbox examples.
Try using CGPANN. for further details you can see my publications in my Research gate profile.
Arbab Masood AhmadFollowing
- Can anyone recommend me some literature concerning live suicidal behaviors and possible prevention?
We are doing a multiple-case study of youth live suicidal behaviors online and trying to figure out possible prevention. For live suicidal behaviors, we mean suicidal behaviors on social media, posting the suicide process lively. Anybody could recommend us some relevant reference? If you could provide some articles on mutilple-case study, that would be perfect!
I have done some research on this, articles below, and it is of course a complex area. It sounds like you are looking for case studies, examples of what suicidal people posted and how. Some of the above articles are decent, some not so much. Here is a newspaper report on this: Grenoble, R. (2012, October 11). Amanda Todd: Bullied Canadian teen commits suicide after prolonged battle online and in school. The Huffington Post,. Retrieved from http://www.huffingtonpost.com/2012/10/11/amanda-todd-suicide-bullying_n_1959909.html?utm_hp_ref=mostpopular
And a more academic approach to the same topic:
Lester, D., McSwain, S., & Gunn Iii, J. F. (2013). Suicide and the internet: The case of Amanda Todd. International Journal of Emergency Mental Health, 15(2), 179-180.
However, if you are looking for a more systematic analysis, those are more difficult to find as there are many limitations in this kind of research.
Not sure if you can obtain the full text, maybe contact the lead author, but this is an interesting perspective on the situation:
Li, T. M. H., Ng, B. C. M., Chau, M., Wong, P. W. C., & Yip, P. S. F. (2013) Collective intelligence for suicide surveillance in web forums. Pacific Asia Workshop on Intelligence and Security Informatics, PAISI 2013: Vol. 8039 LNCS (pp. 29-37). Beijing.
Another interesting study: McSwain, S., Lester, D., & Gunn Iii, J. F. (2012). Warning signs for suicide in internet forums. Psychological Reports, 111(1), 186-188. doi: 10.2466/12.13.pr0.111.4.186-188
Here is some of my work.
A summary and discussion of some of the themes you appear interested in:
Harris, K. M. (2015). Life vs. death: The suicidal mind online. In E. Aboujaoude & V. Starcevic (Eds.), Mental Health in the Digital Age: Grave Dangers, Great Promise (pp. 135-151). New York, NY: Oxford University Press.
A study on suicidal individual's online suicidal behaviors, both positive and negative:
Harris, K. M., McLean, J. P., & Sheffield, J. (2009). Examining suicide-risk individuals who go online for suicide-related purposes. Archives of Suicide Research, 13(3), 264-276. doi: 10.1080/13811110903044419
A study on more everyday online activities that suicidal people engage in:
Harris, K. M., McLean, J. P., & Sheffield, J. (2014). Suicidal and online: How do online behaviors inform us of this high-risk population? Death Studies, 38(6), 387-394. doi: 10.1080/07481187.2013.768313
A more in-depth exploration of the suicidal mind, including online behavior:
Harris, K. M., McLean, J. P., Sheffield, J., & Jobes, D. (2010). The internal suicide debate hypothesis: Exploring the life versus death struggle. Suicide and Life-Threatening Behavior, 40(2), 181-192. doi: 10.1521/suli.2010.40.2.181
You can find these on my profile here. Also, please get in touch, would be very happy to discuss your project further!
- Which drugs can activate the Hypoxia Inducible Factors in kidney?
I am looking for the drugs that can activate Hypoxia Inducible Factors in kidney. Usually used drugs are prolyl hydroxylase inhibitors. I shall choose one of them to use in mice with a long period about 6 weeks, because I need to explore its role in chronic kidney disease animal models, instead of acute kidney disease. Thanks a lot.
Thank you, Dr. Jelkmann. We don't care about hemoglobin.Following
- How a correlation is made for a specific problem?
Ok, if we have a 2D steady state thermal conduction problem,( constant heat flux , convection , insulation Boundary Conditions), and we do not want to solve a specific problem but have a general statement on how one variable changes with regard to other variables (= a correlation), How is it possible to do so?
*Should we non-dimensionalize it? (but then what..)
*Should we change EVERY variable one by one, while others being kept constant ?
T=T(x,y,q",h,x0,y0,K,T_inf)--------> T*=T*(x*,y*, h*,q*) ?Following
- Why isn’t nutrition a bigger part of conventional medical school education? Diet is arguably the single most important preventive measure for healthy aging because it affects the functioning of every organ in the body and is a factor both in the development of disease and in recovery.
Drugs are patented, giving a financial incentive to pay for research, and to pay for publication in open access journals. Paper journals have an incentive to publish research on drugs, so that they can make money from selling offprints to the drug companies. University staff need to publish research if they wish to be promoted. They need research funds and journals that will publish their work. So doctors interested in nutrition are handicapped. Therefore medical students only learn, at best, mostly simplistic nutrition, and at worst, false nutrition.
Money is available for reversing disease, rather than for preventing it in the first place. Maintaining health is invisible, while treating disease looks good.
Manufacturers of processed food spend much money advertising their products, providing teaching materials, and influencing food policy.
Double blind studies have their limitations. They cost a lot of money, and he who pays the piper tends to choose the tune. Also you cannot do a double blind study on parachutes, for example. Long-term double blind studies on real foods are not possible.
Double blind studies on nutritional supplements save money by using cheap supplements, like synthetic vitamin E or synthetic beta carotene. Synthetic vitamin E contains only 1 of the 8 forms of the vitamin, and the mirror image of that one. It blocks uptake of the other 7. Pure synthetic beta carotene blocks uptake of the many other important carotenoids. Doses chosen can be too high for safety, as in the ISIS study, or too low for effectiveness. Medically trained researchers do not necessarily know enough nutrition to design reliable studies.
Studies may use inappropriate combinations of nutrients. For example, the Linxian cataract study used retinal and zinc in one group, vitamins B2 and B3 in another, and vitamin C and molybdenum in a third. Yet vitamin B3 and zinc cooperate to process retinol and retinal. Vitamin B2 and molybdenum cooperate in various processes. Vitamin B6 was used with progestogen and no vitamin B2 in the Dalton and Dalton paper. Yet progestogen depletes B2, and B2 is needed to activate B6.
Because people should not be taking a drug before it is prescribed, it can be prescribed without looking at intake from other sources, However it makes no sense to prescribe high dose calcium to those already on high calcium diets. Excessive total calcium intake blocks magnesium absorption, which may lead to death from heart disease. Supplementing selenium in someone who loves Brazil nuts can lead to dangerously high selenium levels. Giving vitamin A to someone who eats lots of liver can lead to death of infants, premature cessation of growth of long bones, or pseudo tumour cerebri, a feeling of pressure in the head. Prescribing a multivitamin to someone already taking an antioxidant and cod liver oil can result in harmful polypharmacy, with excessive intake of selenium and vitamin A.
I suggest we start the training of doctors in the community, where they could study disease prevention, learning about nutrition, sleep, exercise, sanitation, hygiene and pollution. Only when they understood about health would they go into hospitals, to find out about what to do when health breaks down. They could learn to use nutrition as their primary tool, and toxic drugs only when really needed.Following
- How should I explain and justify a jacobians matrix for which the eigenvalues are equal to zero?Is my system of equations stiff or not?
in my equations system ..i write jacobians matrix and i calculate the eigenvalue of my jacobian matrix.. but the two eigenvalues from three eigenvalues are zero and my stiifness ratio is infinity .
Zero real part could mean unstable system, but may require further investigation, like through phase plane analysis. see http://people.uleth.ca/~roussel/nld/stability.pdfFollowing
- Can someone help me with my moderation study conceptual framework model?
I have some confusion about my moderation study and hope someone can brighten my way. So here it is (i also attached picture pdf file to give you more explanation about my question)
So basically, my research has 4 variables.
- 1 Predictor Variable
- 1 Criterion Variable
- 2 Moderator Variable
The purpose of my research is to find out which among the two variable is a better/stronger moderator variable. What i mean by better/stronger is which one of them have stronger buffering effect to the relationship between the predictor variable and criterion variable.
So upon working on this topic, i face a confusion regarding the model (conceptual framework) of this research. Which could lead to different number of sample size.
If you can download the pdf file in this post, you can see that there are two model there. "Model A" is the model that the panel proposed to me. They want to compare which among this moderator variable is the better variable.
My question is, do model B is the same with model A? "Model B" is the model that i got from previous research.
I ask this because this can lead to the different determination of my sample number. Let say i use the Model A, and participant determination formula from Green (1991) which is 50+8k (k = number of predictor variable). Do i have to find separate participant to each moderation equation, so 74 persons for equation A (social support as a moderator) and 74 persons for equation B (Perceived Financial Security as a moderator). Each participant will be given 3 questionnaires (different questionnaire for the moderator variable). After its done, i will calculate and compare the result. The total [participant will be 148.
I use Model B where the moderation taken together in one equation and have the same participants. The number of participant will be 82. Each participant will be given 4 questionnaire to complete.
So that is my question, i am so sorry if this question kind of "uneducated" but i am really new to this moderation study and wanting to learn.
Thank you and hope somebody could help me. Thanks
Hello, I prefer model B because you can eliminate variance among respondents if you ask two different groups of respondents. Anyhow, when you test the moderating effect you will do it sequentially just like model A. I share a tutorial link from Statwiki that you can watch the video. It is a good resource for my research.
- Can anyone suggest why there is no chromatin smear after sonication?
HI, I am trying to perform ChIP on low cell numbers (1x106, 200.000) and after sonication and reverse crosslinking to check the chromatin, I see a very sharp band around 1000 bp but nearly no visible smear. Did anybody ever experience the same problem?
thank you for your help. I found that the problem was caused by the SDS. I reduced the amount of SDS and now everything works fine. I thought I just quickly report it here, if someone has faced the same problems and is searching for answers like I was :)Following
- Which will be the best surgery for III degree UV prolapse in a 30 year old woman who has completed her family and has a big hypertrophied cervix? .
If "Weakness, damage to nerve supply of levator ani muscle are the main culprit." how do you explain a >90% cure rate for apical uterine prolapse by repair of only cardinal and uterosacral ligaments?Following
- Which method of laparoscopic cholecystectomy (American or French) has advantages in terms of ergonomics of operation performance?
What do you think, which method of four-port laparoscopic cholecystectomy (American or French) has advantages in terms of ergonomic conditions of access?
Hello to everyone:
I have to agree with most of the previous answers to this topic. I too had most of the times performed multi-port laparoscopic cholecystectomy using the American position (more than 5,000 cases). I use one 12-mm umbilical port (Hasson) and three 5-mm subcostal ports.
However, the reason I prefer the American position is Operating Room time economy, and obviously the cost for it.
My experience with the French position comes from my early Nissen operations and my early bariatric cases. I started doing my first 25 gastric bypass cases standing between the patient's legs. And then I noticed that just positioning and fixing the patient to the table took us in average 15 to 25 min. Therefore I started to operate all my bariatric cases standing on the right side of my patients. From there I switched all my Nissen fundoplications also to an American position and decreased my operating time by 20 min at least.
Now, in regards to laparoscopic cholecystectomy, as I said before, I do almost all of them in the American position. However, when I have to do a Nissen and a Cholecystectomy in the same patient, I start operating from the patient's right side and my 5 ports distribution is as follows:
1) 12-mm umbilical port (Hasson)
2) 5-mm right subcostal port (just lateral to the mid-clavicular line)
3) 5-mm subxyphoidal port
4) 5-mm left subcostal port (just lateral to the mid-clavicular line)
5) 5-mm left subcostal port (between the anterior and middle axillary lines)
Then, when I finished the Nissen, I shift my position to the patient's left side and use the umbilical port for the camera man, the subxiphoidal port for my assistant in retracting the gallbladder fundus upwards and I use the two paramedian ports for most of the operation - like the French technique. Nonetheless, my patient is in the American position. Taking advantage of the best of both worlds.
I hope that this rather elaborated answer contributes to this well nourished forum.Following
- Could someone tell me the name of this copepod species?
Primarily, I am now doing a phylogeography of tide-pool copepod (Tigriopus japonicus) in Taiwan with my colleagues. As far as I know only copepod from Tigriopus spp. could survive in the tide-pools but during my sampling in Penghu Islands and Green Island, we also found this species in the tide-pools that we are not sure that it is Tigriopus japonicus.
The first photo was the unknown copepod species and the second was the Tigriopus japonicus.
Hi! S/o Cyclopoida, fam. Oncaeidae, m.b., Oncaea sp. For identification need more detail and size.
Sorry! It is my vistake!Following
- Dear, Citations to my articles are not updated automatically , does anyone know why?
Citations to my articles are not updated automatically, does anyone know why? please.Following
- How can I design hydrothermal experiments ? And, how to use the thermodynamic modeling ?
Thermodynamic modeling can be used to design a process to be thermodynamically favored using fundamental principles instead of the Edisonian methods. BUT, how can I realize it?Following
- Any one have the idea about state of the art algorithm about features extraction matching in 3D point Cloud?
i am searching the some recent paper about topic:
"state of the art: features extraction and matching algorithm in 3D point Cloud"
Have you seen PCL - Point Cloud Library (PCL)?Following
- Is there an alternative way for Weibull distribution method in wind power potential?
Is there an alternative way for Weibull distribution method in wind power potential?Following
- Are toxicity studies required for functional foods?
we are developing a functional food , so would like to know whether sub- chronic toxicity study is required.
The best place to find out is to contact your government regulating body such as FDA in US and TGA in Australia... Check with them before you proceed any further as every country has slightly different regulations.Following
- How can I protect a primary hydroxy group with cbz group having already a NHCbz group in my molecule?
how can I protect a primary hydroxy group with cbz group having already a NHCbz group in my molecule but that should not get diprotected?Following
- Can anyone help me please?
I'm looking for Lloyd A. T., Todd N. B. 1989 Domestic cat gene frequencies: a catalogue and bibliography // Tetrahedron Publications, Newcastle Upon TyneFollowing
- What is the input port power of a cell phone antenna?
What is the input port power of a cell phone antenna?Following