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Tue, 1 May 2007 15:56:13 -0500 - Biodot System Tools: BioChip, Biosensor, Lateral Flow
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Tue, 1 May 2007 15:47:54 -0500 - PLS HELP ASAP: Assaying transient VSCGs w/ voltage sensitive dyes
Hello All, This is my first posting & I hope some of you out there can help. I would like to perform some studies of the NaV family using the voltage-sensitive dyes. Unfortunately I do not have easy access to a rig; I do have access to an FDSS (sim. to the TETRA). I have seen several papers using these VS dyes to measure response w/ recombinant NaVs (293 or CHO) in the presence of veratridine with and without ATX2 & demonstrating response (Benjamin 2005, Felix 2004, posters at conferences (Astra Zeneca 2005, Ionix 2004) 1) Isn't the channel in an inactivated state, considering the RMP of the host cell is higher than the activation threshold (CHO=0- -10 & HEK= -20 - -40mV)? 2) To my knowledge veratridine binds to the open state, so I do not understand how veratridine can work if the channel is indeed inactivated. Can someone explain this? 3) Is it that these groups are using a stable, or one chosen w/ a lower RMP? Considering my results w/ ligand gated channels, etc. the expression levels should not be an issue. 4) Beta Subunits: Is it better or worse to express them w/ a flourometric readout? I greatly appreciate your help, and hopefully can return the kindness in the future. Vicky :-)
Tue, 1 May 2007 15:29:39 -0500 - Pymol: HighResolution Raytracing on large Protein Complexes?
[quote=ryan_m]If you were doing this on Linux you could use "nice" to limit the resources that Pymol uses. My guess is even without niceing the job you would not crash the system but rather just use swap memory until the job completes.[/quote] I also tried to run the whole thing on Suse Linux 10.1 64bit. Pymol crashed as well, the application terminated without any comment... I have only very limited linux skills, but I will try to start pymol with increased "nice" increment and post my experience here... ;-) Thanks for all the good advice! Perhaps it's just a little too keen to render a whole virus capsid...? Should I wait for quantum computers? :)
Tue, 1 May 2007 07:53:49 -0500 - Pymol: HighResolution Raytracing on large Protein Complexes?
[quote=bgood]Hi, "You may try to decrease the value of hash_max, which will reduce the memory requirement for ray tracing, at the expenso of longer rendering time. E.g: set hash_max, 50 " [/quote] I tried with the has_max set to 50 and 20, both of this didn't work as well... neither on linux.
Tue, 1 May 2007 07:49:41 -0500 - Pymol: HighResolution Raytracing on large Protein Complexes?
[quote=frasermoss]so for your 1000 x 1000 you should type in the GUI png yourproteinname, dpi=1000 I have saved 1200 dpi images this way on my laptop running Windows XP, a 1.73GHz Pentium M with 1.5 GB RAM without hassle. [/quote] This is a nice command, but it does not alter the dimension of a picture, e.g. if the picture on your screen is 700x900px, the png command does not change that. It just makes the picture smaller with higher resolution or bigger with lower resolution. What I would like to do, is to really increase the number of pixels... so I can print it for instance as DIN A3 format.
Tue, 1 May 2007 07:42:26 -0500 - scFv
In which order are VL and VH more likely to give a good scFv, VH-VL or VL-VH (N- to C-terminus)?
Tue, 1 May 2007 06:41:02 -0500 - Our 3T3-L1 don't differentiate anymore!!!
They taught me that as soon as we want them to proliferate (=fibroblasts) we never have to leave them to become confluent. But once we seed them to start with our experiment, we allow them to be confluent and wait 24-48hrs before adding the differentiation cocktail (dex+IBMX+insulin or Indomethacin+insulin) i.e. day 0 of differentiation. The meaning is that once they are confluent they, let's say, walk outside the cell cycle and start by themselves the differentiation program: we add the cocktail just to synchronize their differentiation and have a condition of a population of homogeneous adipocytes...if you know what I mean... Moreover, believe me, it worked out very well until the beginning of 2007. Anyway I will try to differentiate earlier (the day of confluence) the next stock I'm going to thaw... I will let you know! Thank you SanDiablo! Meanwhile, other suggestions are welcome!! Silvia
Tue, 1 May 2007 05:51:33 -0500 - Our 3T3-L1 don't differentiate anymore!!!
Typically, cells lose their ability to differentiate if they EVER over grow, and you cannot recover this ability. You'll have to return to the earliest stock of cells that you have and be careful that they never become confluent. If you have to plate for a long weekend, always plate several different amounts, and then only continue the cultures that do not become confluent. Good luck.
Mon, 30 Apr 2007 21:57:10 -0500 - 3T3-L1 cells die in culture
Hi all, I've just posted a similar topic just because I didn't understand which is the solution that Aspeedasai found.... Can you explain me? Thank you Silvia
Mon, 30 Apr 2007 15:34:11 -0500 - Our 3T3-L1 don't differentiate anymore!!!
It's been two months that our pre-adipocytes don't differentiate anymore. We thought that this new lot had problems due to ATCC handling. We changed the differentiation stock solutions (dexamethasone, IBMX or Indomethacin), plates, everything but the media and calf serum. Also a vial from the old stock gave us the same problems: they easily proliferate but fail to differentiate, i.e. they slowly accumulate lipid droplets and they become older compared to the days of differentiation and the degree of lipid accumulation. I found a topic started by Aspeedasai but I cannot communicate with him (I'm new of this forum) so I'm now asking to all the community. Can somene help me? Thanks in advance, Regards from Italy!
Mon, 30 Apr 2007 11:01:31 -0500 - sybr safe
Has anyone used sybr safe with a DGGE gel yet? If so what is the best method of staining...adding to the gel or soaking after running? Thanks for your help!
Mon, 30 Apr 2007 07:13:06 -0500 - Gel Zymography
Hello, As a part of my ongoing experiment, I run Gelatin-SDS-PAGE zymography for semi quantative coparitive study of MMP 9 and 2 on my samples. The samples are unconditioned (not treated with PMA: phorbyl myristate acetate) cell culture supernatant which include 5% FBS. The problem I'm encountering is that eventhough my standard marker is giving me a bright sharp band, my samples are giving me smears. Since more or less this is a specific technique for gelatinases, and I am aware of the presence of Latent (proenzme) gelatinase in the sample, I would thank if anyone could advise me what to do to emit the smear resulting sharper bands.
Mon, 30 Apr 2007 07:07:24 -0500 - Error Bars and significance
I agree. Apparently my PI sent out this article to us saying it to be one of the best for biologists who have to deal with statistics. Should say a must.
Sun, 29 Apr 2007 13:55:39 -0500 - denaturate a lipase
[quote=stud.polyt.bio]i'm a BA student from denmark and i'm trying to synthesize a ibuprofen-glucose-esther with a lipase from [i]rhizomucor miehei[/i] as a catalyst. in order to analyze my product i need to stop the reaction - preferably without heating. does anyone know how to denaturate the enzyme without heating the reaction?[/quote] For your solutions, would it work to partition it away with several non-aqueous extractions? Lipases are highly partitioned at lipid interfaces.
Sun, 29 Apr 2007 11:59:03 -0500
