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How does low dose of BMP-2 affect osteogenic differentiation?
Hello. I am having some experiment about differentiation periosteal-derived cells to osteoblasts in 2D culture. I cultured cells in OM (osteogenic medium) and additionaly certain amount of BMP-2 was added in medium.
After 7, 14, 28 days I checked ALP activty and calcium deposition. ALP was increased until 14days, but calcium was not detected until 28 days. Plus I did RT-PCR, but late marker (osteocalcin, osteopontin) was not detected.
Has anyone eperienced about osteogenic undifferentiation, please answer.
Thank you for reading and sorry for my bad english.
Hi. Can you say what was in your OM? There can be few reasons related to the constitutes of OM eg dexamethasone. in human cells osteocalcin expression is decreased by dex. Did you use any dex? Also, what was the cell density at the start of your experiment?Following
Could anyone suggest why I get very little amount of plasmid DNA after extraction from E. coli.?
I already obtained two plasmid that harboring gene synthesis A or B. First is pUC57 harboring gene A (size of insert 1.5 kb). Second is pUC57 harboring gene B (size of insert 1.6 kb). Then, I transform these plasmid to E. coli DH5Alpha (Rubidiub chloride-Chemical competent cell). After get the transformants, I pick single colony of each plasmid, culture in LB+Amp 3 ml, Amp 100 ug/ml, 16 h and extract it by alkaline lysis method by using culture 1.5 ml or 3 ml. Finally, after ethanol precipitation step, I put 30 ul of TE buffer, and Run 2 ul on 0.8% agarose gel.
For pUC57-gene A is get a lot amount of plasmid. But for pUC57-gene B, I get very small amount of plasmid. I very confuse because I transform many time and pick many transformants to extract, but my result still the same. I doubt because difference only insert but the vector backbone still the same.
Could any one suggest me, how to disslove this problem?
If the two genes have different promoters the two genes may be expressed at different rates and the rate of transcription could affect the rate of plasmid replication.Following
What is the difference between Attributable risk (AR) and Population Attributable Risk (PAR), from application point of view?
One has to look at the applications of AR and PAR and distinguish between the two.Following
What happens when gibb's free energy value is positive in Batch mode adsorption studies?
actually i choosed the initial concentration , more than need.. so distribution coefficient became less than 1.. therefore DG0 is positive... i should say that with increasing the temperature, DG0 becames negative.. in mathematical form this is obvious.. because from DG0=DH0 - TDS0 , when T increases, ( with considering to positive values of DH and DS ), DG is going to have negative value and the process will be the spontaneous one..
Thank you in advance..
I am talking about physisorption (no chemical bonds formed), in this case, the adsorption is exothermic (no strong bonds are broken, and a weak bond is formed).
In the case of chemisorption, adsorption can be exo- or endothermic, depending on the reaction (bonds are being continuously broke, so depends on which is stronger).
Is the case of your more than 10 papers with endothermal adsorption chemisorption?Following
How can I model the glad design theoretically in the software like open filters, filmstar, optilayers?
Modelling the design for GLAD
Hi..Sorry..I don't know..Following
Any smart grid simulation software/tools that are avaliablle?
Given the scale and complicity of smart grid network, it is very hard to model the whole system in an accurate model. Instead, simulation may be an effective and efficient way to study and evaluate various scenarios and technologies. I wonder if such smart grid simulation software or tools exist and have widely been used.
Conversation in the attached link might be useful. Thanks.Following
What is the example of IF ring which is not von Neumann regular?
I am trying to find an example of IF ring R (every injective R-module is flat) which is not von Neumann regular (every R-module is flat).
Let R is R Z+Q((x)) then R/J(R) is IF ring which is not von Neumann regularFollowing
How to classify your students through the quality and nature of their questions?
Some students ask smart questions, while others ask stupid, silly, ridiculous, and meaningless ones. Some of them ask wise questions while others ask pointless and aimless ones. Some are interested only in issues that would be most likely asked in the exam. Some ask irrelevant questions to divert attention.
Please provide us with your personal experience and opinion.
We need to classify questions as average questions, good questions and excellent questions. Average questions are those questions usually every student ask and we can answer them directly. For certain questions, we need to explain the solutions with help of some illustrations and explanations. These are good questions. Some questions are really challenging and thought provoking for us and we need further studies and references to answer them.
All students ask average questions, but only excellent students student ask excellent questions. Excellent students can be identified in view of their questions. But I am not sure whether other students can be classified merely in terms of the nature of questions they ask.Following
Is Chalmers' so-called "hard problem" in consciousness real?
In his 2014 book "Consciousness and the Brain: Deciphering How the Brain Codes Our Thoughts" Stanislas Dehaene wrote "Chalmers, a philosopher of the University of Arizona, is famous for introducing a distinction between the easy and the hard problems. The easy problem of consciousness, he argues, consists in explaining the many functions of the brain: how do we recognize a face, a word, or a landscape? How do we extract information form the senses and use it to guide our behavior? How do we generate sentences to describe what we feel?
“Although all these questions are associated with consciousness,” Chalmers argues, “they all concern the objective mechanisms of the cognitive system, and consequently, we have every reason to expect that continued work in cognitive psychology and neuroscience will answer them. By contrast the hard problem is the “question of how physical processes in the brain give rise to subjective experience … the way things feel for the subject. When we see for example, we experience visual sensations, such as that of vivid blue. Or think of the ineffable sound of a distant oboe, the agony of an intense pain, the sparkle of happiness or the meditative quality of a moment lost in thought … It is these phenomena that poses the real mystery of the mind”."
Stanislas Dehaene's opinion is "that Chalmers swapped the labels: it is the “easy” problem that is hard, while the “hard” problem just seems hard because it engages ill-defined intuitions. Once our intuition is educated by cognitive neuroscience and computer simulations, Chalmers’ “hard problem” will evaporate".
Personally, I agree with Stanislas Dehaene's opinion.
As usual, your comment, Marc, is senseless. Statistical moments are first year high school education.Following
How to define interface between two different thickness and different material in DIANA?
I am modeling infilled frame in TNO DIANA. For infill frame joint, i define interface using free edge, but i am confused that it will work or not. Because, when i apply load, load is transferred to wall but not in expected way (i.e. diagonal strut formation) and also force vs displacement response is also not looks good.There is convergence problem also. Can anyone please guide me to solve these issue?
Yes sir, if i define normal stiffness modulus (Kn) and tangential stiffness modulus (Kt) for interface, how can i find the direction of local axis should be in which direction? Can you please explain.Following
Can anyone suggest what is the formula for Dave cluster validity index and also where can i find MATLAB code for Dave's index ?
HI, I am using different clustering algorithms for my project. But now am in need of cluster validity index named Dave for my work. So please any suggest me the MATLAB code for Dave index.
Please go through R. N. Dave, “Validating fuzzy partitions obtained through c-shells clustering,” Pattern Recognition
Letters, vol. 17, pp. 613-623, 1996.Following
What is your opinion about mid-Bronze Age (ca. 3.5-3.0 ka BP) climate condition in Central Europe?
The mid-Bronze Age is a period of interesting cultural processes recorded by archaeologists. But reconstructions of climate conditions ca 3.-3.0 ka BP are different. Do you have an opinion or a paper you'd recommend on palaeoclimatic, paleaohydrologic and palaeoecological conditions of this period?
A most interesting question. A summary paper is available from the Central Italian Alps covering the alitude from 1500m til 2200 m a.s.l. .Ask me for a copy. My email addr. is email@example.com . An archaeological-vegetation-historical paper covering the Bronze age- Early iron age from the same area is in preparation.
How do I improve on the blasting results in order to reduce costs in a quarry?
How do I improve on the blasting results in order to reduce costs in a quarry with too much overburden and heterogeneous limestone bands or seams?
the quarry has discrete limestone pockets and usually mined at break-even ratios. sometimes the cutoff grade is never sustainable for mine operations.
This quarry is in Zambia, chilanga run by Lafarge cement plc.
There is water on the lower bench flow.
I might need more clarity on Geosynthetics. All I know is that they are man-made products that are used to provide stability to terrain. I am wondering how this can be of use in the mining sector especially during blasting!
Looking forward to your favourable response sir. Enjoy your DayFollowing
If a transcription factor (AP2) has an alternative spliced form without the AP2 domain, can its overexpression in a line give stress tolerance?
If a Predicted Transcription factor (AP2) having alternative spliced form (Not given in database) and the overexpresion of that gene shows stress tolerance, is it worth for studying further because main AP2 domain was cut out in the alternative spliced form.
Also, in case of proteins that form complexes, splice forms that lack the functional domain can be regulatory too. Do a search for 'microproteins'.
What is the best reference book for electrochemistry analysis?
e.g. CV, DPV, etc.
Thank you very much Rana, I think the book you have sent me, can help me to better undestand the fascinating word of corrosionFollowing
How do you remove the supernatants from an exosome ultracentrifugation sample without disturbing the pellet?
I often use thick-wall ultracentrifugation tubes to pellet exosomes using SW28 swing rotor. I cannot see the pellet after 100,000xg centrifugation. I try to remove the supernatants as much as possible but I am not sure how to leave the pellet intact. Should I pour the supernatants away and invert the tube on a paper towel then aspirate the rest of the medium from the wall?
I used to use standard ultracentrifugation technique to isolate microvesicles released from stem cells. With this technique, after the first round of ultracentrifugation, the pellet is rarely visible with the naked eyes. However, sometimes, you might see a very thiny tiny white pellet centered down to the bottom of the ultraclear tube. Anyway, for recovering this pellet, we get consistant yield by just aspirating the supernatant carefully, so that you can almost dry out the pellet. Afterwards, the resuspension of the pellet can be tricky sometimes...because of exosomes agglomerates. Likewise, after the second round of ultracentrifugation, you should get a very thin white pellet into the center of the bottom of the tube. Again, I think your pellet can definitely be difficult to see depending on the initial cells amount which you're working with. In our hands, usually this pellet is strongly adherent to the bottom of the tube. No risk to detach it from the tube when you aspirate the supernatant. You just have to make sure you do not directly aspirate the pellet with your Pasteur pipette. Finally, you could try to stain your exosomes or microvesicles just to track them in order to make them visible so that you can localize the colored-pellet on the bottom of your tube. this is an interesting method to visualize your precious pellet !
What could be the origin/source of even carbon number n-Alkenes from C12 to C22 in sedimentary organic matter?
Presence of n-alkenes in sediments have reasonable maturity is questionable from long time. If anyone have clue about origin/source of even carbon number n-Alkenes from C12 to C22 in sediments.
thanks to all, but please note the question is about alkenes, not alkanes.. most of you talked about alkanes..Following
How biosafety measures help us in prevention of virus? which factor or thing is behind it that force us to adapt this type of analysis?
In case of viral diseases experiments in labs we adopt biosafety measures like dress that should cover whole body etc. but question is that how biosafety measures help us in prevention of virus? which factor or thing is behind it that force us to adapt this type of analysis?Following
How do I run a rapidminer process using java file in eclipse?
I want to run the process of rapidminer using java file. I tried many times but could not get the output. Please can you help me for integrating rapidminer and eclipse.
I am also sending my file along with error. Please guide me.
@Ketan Sarvakar..Thank you so much..
It really helped me a lotFollowing
Why does seal resistance fall to less than 50 MOhms when I do whole cell patch clamp?
I am new to electrophysiology in general and I am doing whole cell patch clamp at the moment to investigate the impact of centipede venom on NaV 1.7 expressed by TE671 cells. For some couple of days after learning patch clamps I had everything going on so perfectly with brilliant seals and excellent patches coupled with low leakages ( picture a). However, for some time now my good days are gone as recently I hardly have good patches formed. It appears when I apply negative pressure after the tip of the pipette just touches the cell membrane, the seal resistance increases to somewhat 500-800 MOhms and then decreases to less than 50 MOhms ( picture d) making it impossible measure a sodium current coupled with large leakages (nA). Maybe the pictures may help explain the issue more NB: Pipette solution is 140mM CsCl, 1mM MgCl2, 11mM EGTA, and 5mM HERPES, pH 7.2. We do not fire polish the tips of the pipette in our lab as patch clamps have been achieved in the pass without fire polishing.
Applying the negative pressure following the giga seal formation is the most critical step. During your experiments you will get the experience that how much negative pressure is the best. Please also adjust the amount of positive pressure which is applied through the micropipette. Sometimes a high positive pressure may be harmful for the cell, but sometimes it is necessary to clean the surface of the cell and to produce a good dimple before releasing it. Thus, it very difficult to distinguish which part is not correctly done in your experiments. The comments of Thor and Elise are alsovery critical.Following
What is the average pileup number for good peaks in CHIP-seq data validation?
I'm currently trying to find some good peaks in my CHIP seq data (with the aid of UCSC genome browser) which I could use for data validation (primer design, qPCR) and wondering how many (average) reads a "good" peak should have. They look quite nice when the pileup is around 20-50, but can I trust a peak with about 5-15 reads, too? And what do artifacts look like?
Does anyone have some examples or knows about good help-webpages like tutorials etc.?
I unfortunately do not have experience in validation yet, so I would be happy to get some tips.
To judge whether your experiment was successful, you can also have a look at the efficiency measure (that is, the percentage of uniquely mapped reads that are found within peaks called by MACS). It is an easy to calculate measure of your signal to noise, and it can be obtained for example with Galaxy or bed tools. A reasonable ChIP-Seq experiment should have an efficiency of at least 5%, and really good experiments can go up to 20-40%.
To exclude artefacts, one thing I would recommend is excluding peaks that have high signal in your matched input sample. This can be especially important when using cell line material, but a good idea in any case.
For your QPCR validation, I would suggest that you use a few peak regions with different intensities in your data (e.g. low, medium and high) so that you can see how reliable these are in terms of QPCR enrichment.
Hope that helps! :)
Which tests are preferred for validating tools used in assessing impact of learning modules on knowledge, attitude & skills of undergraduate students?
We are planning to conduct an impact assessment study of gender-integrated modules taught by trained medical faculty to undergraduate medical students, in order to assess their learning in terms of knowledge, attitudes and skills, as compared to students who have studied the standard medical curriculum in the state. A quasi-experimental research design will be used and pre and post-tests will be conducted with experimental and control groups of students.
Can anyone suggest tools (or provide links for tools) which have been used in similar studies? Also, readings on how to carry out validation of tools would be most helpful.
Thanks in advance!
@ Balakrishnan Ramaswamy: Thank you for responding. I'm not able to follow the answer due to its formatting. Could you please share your answer again if possible?Following
Is it normal to observe many bubbles during heating up of a mixture containing oleic acid, 1-octadene and lanthanide acetate solution?
When the temperature is close to 120oC under stirring (I need to heat up to 150oC form lanthanide oleate), suddenly a vast amount of bubble give out like soaps and they eventually disappear. It is a bit strange because I dont observe vast amount of bubbles before. I afraid this was one of the factor affect my nanocrystal synthesis.
Before the existence of the bubbles, the mixture stay a bit turbid and the mixture become clear again after the bubble evaporation. Actually, I suspect the destruction of oleic acid as indicated by the vast amount of bubbles
I had the same problem in the past, while using reagents which absorb oxygen or water vapor. Try to do what John suggests, this usually solves the problem. I am using also oleic acid and I always follow a degassing procedure (in a Schlenk line under vacuum) before using it.Following
How well do riboswitches work in regulating gene expression (on/off)?
I would like to see the riboswitches mediated on/off of a target gene.
These articles will help you.
Anyone familiar with the AM2A oxide collector reagent especially the conditions that must be maintained to have high recovery?
This is used in Copper flotation. I am performing lab tests but there are just a lot of unknown factors like what pH should I use or how much conditioning time I need.Following
Which antibody panel is beter for quantification of CD4+ regulatory T cells?
There are many Treg cell associated markers, like FOXP3, CD127, CD25 etc. Which panel of antibodies do you use in your Lab for Treg cell analyses?
CD4+, CD25 high (top 5%), CD127 low/negative, FoxP3+ cells.You can stain a small proportion of your cells with FoxP3 and if it looks good you, if you need live cells, use only the three other markers. The GFP reporter described in the answer above would be optimal indeed, just be careful when using FACS so that GFP doesn't leak in the channels you use for the rest of the markers.Following
Any suggestions about a drug reaction vs erythroderma?
This is relating to erythrodermic psoriasis , when a patient is extensively red and scaly affecting almost the entire body. How does one differentiate if he has an ongoing flare of his erythroderma with constitutional symptoms and a possibility of an exanthematic drug rash to Vancomycin given for MRSA skin infection? Can this be guessed on a Skin examination?
I am sending you a table about erythroderma and drugs.
Table II: Drugs Associated with Erythroderma
CLASS OF DRUGS
Antibiotics, antimycotics, antimalarials
Para-amino salicylic acid
β2- adrenergic receptor agonist
Mercury and Mercurials
What is a suitable method for PLGA encapsulation of hydrophobic drug?
I am currently working on PLGA nanoparticles and encapsulation of hydrophobic drug. What (among single emulsion, multiple emulsion and nanoprecipitation)is the best method for enhanced encapsulation of hydrophobic drug ?
Dear Marco, could you please elaborate your answer on nanoprecipitation technique!
Usual practice is to add organic phase to aqueous phase right? Would it work the other way? ThanksFollowing
Can anyone recommend some good references for semi-Markov decision processes with practical applications?
I want to examine stochastic systems in manufacturing that exhibit properties of a semi-Markov decision process;and then develop optimization models.Following