I would like to add something about the NanoDrop versus NanoPearl, I don't think that the path length is fixed on this instrument (the NanoDrop) since if I recall well, the instruments sets the path length as it scans through the droplet, hence the characteristic clicky noise that you hear as the handle moves. This instrument requires recalibration every year (or so according to the vendors, and this is of course not cheap) since there is a moving part that can get out of alignment. On the other hand, the NanoPear from Implen as a fixed geometry for the same tiny amount of sample required. Thus you do not have to deal with moving parts and recalibration. We just bought a NanoPearl and I should also mention that this instrument does both things: nano-drop size measurement like the NanoVue AND cuvette size measurements (for OD600 or old school Bradford).
All the best, On Thu, Jun 16, 2011 at 4:57 PM, Shaun Lott <s.l...@auckland.ac.nz> wrote: > Just to add my 2c worth... > > The department here has a couple of nanodrops as a shared facility, one for > DNA/RNA and one for protein. It has been noticeable that over time people > has been getting decreased reliability of measurements on the latter machine > cf cuvette measurements, presumably due to the build-up of protein deposits > over time - so I would say that although it's easier to clean than a > cuvette, the nanodrop is not immune to the problem. The biggest issue I see > with the nanodrop is evaporation of sample. Even here in moist Auckland, > where RH is very often 80%+, taking a series of measurements with the > nanodrop over a period of just a minute or two shows increasing > concentration in the sample. So, for consistent results, one has to be > careful to measure quickly. It's probably fine for comparative measurements, > but as has been observed above, not great for super-accurate values for > biophysics, and I think rather operator dependent. But all our students are > super-careful, right? ;) Worth to note also that ProtParam calculates > extinction coefficients based on Gill & von Hippel, (Gill, S.C. and von > Hippel, P.H. (1989) Calculation of protein extinction coefficients from > amino acid sequence data. Anal. Biochem. 182:319-326) who claim accuracy of > ~5% for 'normal globular' proteins without extra chromophores. Whilst on > this subject, I would put in a plug for the good old BCA (aka Pierce) assay > for protein concentration. It's a little slow, but gets away from sequence > dependency somewhat as it is primarily dependent on the peptide backbone > rather than sidechains and works well in micro-titre plates etc. It is > certainly very superior to Bradford. (Smith, P.K., et al. (1985). > "Measurement of protein using bicinchoninic acid". Anal. Biochem. 150 (1): > 76–85. doi:10.1016/0003-2697(85)90442-7). > > cheers > > Shaun > -- Pascal F. Egea, PhD Assistant Professor UCLA, David Geffen School of Medicine Department of Biological Chemistry 314 Biomedical Sciences Research Building office (310)-983-3515 lab (310)-983-3516 email pe...@mednet.ucla.edu
