here's a run-through of how to set up the perkin elmer Envision plate reader.
first step power on the equipment: to do this you first turn on the computer which will be
at the back once the computer's powered on and the windows program has started then you turn on the
actual plate reader once the plate reader is on you can go ahead and start the envision program
give it a minute you may be doing some filter checks before it starts up so now this brings you
to the programs that you can run you can do absorbance reading, you can do fluorescent read
you can do luminescent reading in some models you can also do alphascreen reads these new specific
detectors that you would have to add on to the instrument the inspire when it's given to you
doesn't have it so say you are doing an absorbance based plate read what you can do is copy one of
the existing protocols that they've given you so it to do that you just highlight it and then
you have to go to create protocols and in create protocols you can see here that you've got a copy
selected so you can now copy that and if you can see it's popped to here as copy and so now you've
got a copy but you don't... it's not going to be for a bradford assay probably it's going to be
an absorbance read for any other... for some other assay so then what you can do is change the name
so if you come into the settings you can actually get this pop-up screen keyboard where you can say
well you know call it whatever you want you can use the keyboard or can use it on screen
so protocol for absorbance
reads and you can just go okay and now it's named it and you can put instructions here so let's say
this one the read it's not at 595nm but you're doing an absorbance root at 405 nanometers then
and it's not a protein concentration but say you're doing some sort of cytokine measurement
so let's see yeah just name it as you need
cytokine measurement and now it's ready for your specific
assay so what we do is we specifically we copy something that is an absorbance based methodology
and then we name it for ourselves and of course we want to change the specific absorbance so if we're
doing absorbance and we say now we're doing 405 nanometers then here we can change the wavelength
that we want to measure up and these are the heights of the plates that you'll be using you
can usually leave it on the default unless you're using some specialized plate specific plate and
then you need you can select what the kind of reads that you want so you can do a single and
it's doing simple absorbance but if you want like a well scan then you need to bring it over
so you to press this then you can bring it over and you can let it know that it's an absorbance
read and it will bring it here now so you see it's got absorbance and it's got a well
scanned but we don't if you don't want it and we don't want it you can just get rid of it now
so it's got this measurement c we don't want that we'll get rid of that as well but yeah if it's not
a single measurement you can pick whatever measurement maybe it's a spectrum scan and
on fly then to accept it you just press the arrow and then you can add other components if you want
to delay if you want a temperature if you want to shake so say i want it to shake just before
it reads i can bring that here and i can move it up because i want the shake before the read
and then you can specify how long you wanted to shake it for say i just want the shape to be for
you know three seconds and then it reads it then that's where you set it so that's what
you would do and so to save this now sorry then you've got also your outputs and your outputs
here there's an initial plate measurement that it will put out but if you also want like if
you have a standard curve for your measurement that you would have liked so back here if you
know that certain well so in the plate certain wells we're going to be seeing measurements for
your standards so let's mark these as uh standards and normally you do to replicate so
this one's doing single kits but let's just go ahead and delete it so we can instead of single
case we can now acknowledge that we want replicate so this is where you can change your replicate and
the sample because we deleted it we're actually starting back from one it thinks that it's got it
here starting from 13. so here when you click on it it brings the pop-up here so you just change it
here and now it's at one and then if i select the wells here we go standard one standing to standard
three standard four standard five and so here you can actually go ahead and with the standards
enter the concentrations and the way you do that is when you're in these calculations area so if
you come to calculations for the standards we need to bring it across that we've got standards
and if you hit edit then you can put in the concentration so it was two thousand
okay so i'm using the keyboard it's not working so just use the on-screen keyboard
so 2000 picograms (pg) and let's say you're doing a one in two dilution so the next is a thousand
you know whatever your standards are in you just go ahead and fill it all in
so here you can see i've filled up all the concentrations and then you can just go back to
calculations and so now we know we the instrument will be able to calculate the standard curve and
then you can also select the kind of curve fitting that you want for your standard curve so here
we come back to edit the curve fitting types we have linear regression we have spleen we have
4PL (4 parameter Logistic) typically i tend to use a 4PL but it is dependent on the type of response
you're actually the readout is actually producing so let's say we're going for a 4PL fit then
because we've now got a setup where this plate we've got standard curves on it and we've told
it what the concentrations are and we've told you what fit it is when we now go to output
in addition to the plate we have to tell it that we also want the calculated
concentration if you don't do it it's not going to show it so you have to add it here
and now you've got an output and so you'll be able to look at your file settings .csv
if you didn't want that file setting then you change it and it shows you the wells
and that's now you've now set up it's calling a copy of bradford protein assay but don't
forget that we changed to the name so now if i go anywhere if i click anywhere you'll tell me
do you want to save changes and we'll say yes and so now we have that purple that we've just created
based on an original one but it's customized and fitted for the one that we're actually doing
so now let's say we're done we're going to run the plate then while you select it you press run and
you've inserted your plate and it's going to start on the screen here to start reading your plate
and so you'll be able to see it's doing the shake and it's now going to start reading the absorbance
at 405 nanometers and it's going to give you this is just a test run so you can see it's just giving
you all zeros but if you actually have samples in there it would give you the readouts for that
mate so when you're running a plate when you're in the run protocols if you just go to run protocols
it'll start reading assuming that the plate is in there but if you haven't placed your
plate you need to come to load plate here and that's when it will then give you the plate
holder to be able to actually start the ring so it's not immediately obvious the plate won't come
out when you go to run protocol until you hit the load plate and that's when you will get it
so you've read your plate now and you want to access your results the results will pop up
automatically you have to wait for that but say you are trying to access the previous results
then you would go to show results and when you go to show results you want to be able to
access that particular results associated so you highlight the protocol that you ran it under
and then show results and here you'll be able to export the result in a format that you need
and so if i go here to file settings this is csv but say there was a graph that i wanted to export
as a graph then you would use the mht format because that's what the visual representation
is if you use a csv then in excel you'll be able to see your files so to actually access your
file to copy into your particular usb if you're not networked
you need to come and press the windows key once you press the windows key it will give you these
additional icons which you weren't able to see before so then i can press my file
folder file and here i can go to uh desktop which is where the the inspire folder is
inspire manager inspire data so you go to inspire data and here it creates a folder
with the dates of the 25th of the fourth and you'll be able to open it and access that file
now copy it and because you would have placed your usb you can now put your file in there
before you go i've shown you how to create protocols so say you didn't want to create
a protocol but and there was an exact protocol and you just wanted to edit it
then you could also do that so in run protocols you've got all of this say i want a fluorescent
protocol so here i can just go and i'm keeping it i'm not making a copy then you could just press
edit and then you can go ahead and in this press edit again to change specific things and once
again you can come into the heading the settings to change the name you can go into the measurement
to specify what kind of scan you know if you want any shakes or delays or temperatures requirements
and change the you know excitation and emission because this one is now fluorescence read not
just an absorbance read and by just clicking into any other one it will ask you to save and
if you want to save it then go ahead and save it so great so let's go back to now exiting the unit
to close down it's a little bit unintuitive to close down you actually need to go to
settings once you finish your run you go into settings and then it says exit to windows
and so this is where you will be able to go and exit the software we'll go to settings
i'll just say no because i don't want to save it so then we go to exit to windows
and this is how we close the software so you can do only the inspire manager or if you don't
need to use any of the functionalities you can just close whole perkinelma
expire software and go okay and so you just wait for it to shut itself down and once it
shut itself down and then you can turn off the actual computer just by pressing the button