i'm working as a diy bio. I'm finding a way to create a microglass capillary for picking up single cells. I see this video onyoutube and would like to know what is the minimum I/O diameter ifI pull a capillary like this. Can it be 10-50

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Jean KeelingLv2

27 Jun 2019

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I'm developing a cell-based assay in 96-well plates thatrequires adherent cells, as they need to be washed at least twiceduring the protocol. I'm using in-house strains of HT1080 cells(some overexpressing a certain protein of interest) thatunfortunately have been selected for suspension growth inchemically-defined media (50/50 CD-CHO/CD-293). I can convince themto adhere by culturing in DMEM with 10% FBS and growing them inCELLBind flasks, but they still tend to pull away if there is verymuch shearing force, such as during the wash steps, or when they'rein PBS during the assay wash and incubation steps (maybe 2 hourstotal for now, I'm still optimizing that).

Unfortunately, the parameters of my experiments don't allow meto keep the cells in serum-containing media during the assay, as itwould interfere with the uptake of my drug. Like I've said, when inPBS they start detaching, and I would assume the same in HBSS,which I don't really want to use anyway due to its low bufferingcapacity in CO2 incubators.

Might DMEM alone (without FBS) provide a little more impetus tostay attached during the assay? What would the effect be ofincreasing the amount of FBS to say 15 or 20% during culturing -would that promote stronger attachment? I'm a little hesitant tocoat the wells with poly-D-lysine, as my readout is on afluorescent imager, and I'm worried it would substantially increasebackground. I don't really have enough time to re-select the cellsfor adherent growth (although if I'd known about these issues frommy predecessor 3 or 4 months ago I would have started rightaway...), so are there any other tricks I can try in the meantime?Are there any media supplements I could use to promote adherinexpression or something like that? Any tips or advice at all wouldbe appreciated.

I'm having trouble finding a method to find exons in theoriginal DNA sequence used to create the mRNA, even given thesequence of the mRNA, as I cannot find a way to reliably identifythe beginning and end of introns and exons. Is there an algorithmor method that I can use to do this? What information am I missingto be able to find the beginning and end of exons and introns?

Thanks in advance for answers.

First of all I am not endorsing Intelligent Design (Wikipedialink); I'm asking this because I (someone who does not have abackground in biology, organic chemistry, or philosophy) got into aconversation with someone who does endorse it, and I'm trying tosee his point of view as far as rationality can allow. Second ofall, I apologize for any vagueness; Intelligent Design isn't wellenough defined as a theory for me to help that. I also apologize ifthis is rehashing content already plentiful on the Internet, butfor my purposes I can't simply depend on the likes of Wikipedia orTalk Origins, so what I'm looking for here instead is to take theopposite approach and see, based on your informed minds, if therecould possibly be any reasonable likelihood of an ideal ID theorybeing adopted.

I'm looking for the most favorable consideration for what IDmight be if it were best developed as a viable hypothesis or theoryto get an idea of how far such a discussion is worth taking. Anunbiased, informed Devil's advocate, if you will.

My cursory investigation of what Stephen Meyers (video link) andMichael Behe (video link) say (correct me if I'm wrong) seems to bethat either the first cell was likely designed intelligentlyjudging by the complex code of DNA and that evolution took coursefrom there, or that the first cells were made and many instances ofevolution were from some kind of intelligently guided mutations aswas needed to make "irreducibly complex" cells or body functions(whether the guidance stops after the Cambrian or continued even tothe point of making different types of new bacteria and differenttypes of apes/humans or whether every single mutation and adaptiveability is being guided, I'm not sure if they're at any consensus,and that's why I don't know if I should stop my question atabiogenesis or include evolution). The person I spoke with alsobrought up some specific claims (everything looks designed, DNA hasordered, complex information, blood clotting is irreduciblycomplex, ID can and has make predictions like decades ago that"Junk DNA" wasn't junk) that I don't need you to debunk (I haveGoogle).

But if there could be any possible support or truth to any ofthe above, I would appreciate knowing about that. (Again, Devil'sadvocate so I can be informed and understand where the points comefrom.) Reading an ID book alone, it would be in places hard to knowwhere they differ from actual scientific conclusions. After all,when someone who seems to know what they're talking about and canexplain something in technical detail says that something isimpossible or extremely unlikely, it's hard for me to know why theywould be wrong (are they ignoring other possibilities?misrepresenting facts? are they really right and just getting thecold shoulder from an atheistic scientific community because itimplies a deity?). So I focus on mainstream sources, even if theydon't bother entertaining ID and leave me ignorant of its possiblevirtues. And thus I'm hoping someone from this informed communitymight enlighten me to what those possible virtues might be.

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i'm working as a diy bio. I'm finding a way to create a microglass capillary for picking up single cells. I see this video onyoutube and would like to know what is the minimum I/O diameter ifI pull a capillary like this. Can it be 10-50

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