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Steady-state flow refers to the condition where the fluid properties at a point in the system do not change over time.
Laminar flow means the flow is smooth with layers (or lamina) of fluid sliding smoothly past each other (i.e. there is no convective mixing).
Steady state laminar flow is therefore the maintenance of laminar flow with constant properties.
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You will always be measuring the average hydrodynamic radius (Rh) of all labelled species, so you will see the hydrodynamic radius of the average species rather than a breakdown of the proportion of individual species present. You will not be able to get a representation of eg % monomer, dimer, trimer.
The reagent cartridges are currently supplied with ultrapure (Type I) water in the analyte or auxiliary fluid stream.
The microfluidic chips are made from injection molded COC — this allows excellent reproducibility between batches.
There may be a small amount of sample loss associated with “stickiness” of proteins, which is sample dependent. Our chips are coated to minimize protein adhesion and reduce this risk.
The chips are made of COC (cyclic olefin copolymer) and are manufactured by injection molding to achieve high reproducibility.
Any difference in viscosity between the sample and the aux buffer could impact the detected hydrodynamic radius by changing the rate of diffusion of your protein. We are in the process of studying the impact of typical viscogens on measurement accuracy.
If the smaller peptide was labelled with a fluorescent label the size change between the small unbound peptide an the complex would be noticable allowing for determination of the binding affinity.
The layout of the disposable chips is fixed due do the prohibitive cost of a new injection mold tool.
We do have chip prototyping capabilities however, and we’re always open to discussing potentially novel and innovative applications of our technology. Please contact us if you have a specific requirement and we will be happy to discuss.
Properly citing the instruments used in your work ensures clarity and transparency.
When referencing a specific instrument ensure the name is capitalized and followed by the manufacturer in brackets – for example;
When referencing the overall technique please state; microfluidic diffusional sizing (MDS). This ensures that readers can quickly identify the specific technique used.
If you require imagery to show how our instruments and MDS works here are some diagrams to how they work.
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Hydrodynamic, or Stokes, radius of a particle is the radius of a hard sphere that diffuses at the same rate as that particle. When we are considering proteins, they are of course not hard particles, but complex folds of varying compactness and shape. The hydrodynamic radius of a peptide chain can thus vary depending on its folding state. For this reason, when converting hydrodynamic radius to molecular weight we can provide approximate boundaries corresponding to a fully folded, globular peptide in its most compact state, and a fully unfolded chain that has the least compact state.
For the fully folded state the volume of the protein scales with the molecular weight, leading to a relationship of:
Rh ∝ MW1/3
while for unfolded proteins the relationship is approximately:
Rh ∝ MW0.6
The empirically determined curves illustrating these relationships are shown below. The data used for the folded protein series was the collated experimental diffusion measurements in Tyn & Gusek (Prediction of Diffusion Coefficients of Proteins, Biotechnol. Bioeng. 35, 327). For unfolded proteins data from water soluble polymers (polyethyleneglycol, dextran and polyvinylpyrrolidone) in Armstrong, Wenby, Meiselman and Fisher (The Hydrodynamic Radii of Macromolecules and Their Effect on Red Blood Cell Aggregation, Biophys. J., 87, 4259) was used.
We’ve created a converter to enable you to make this conversion for both folded and unfolded proteins—from hydrodynamic radius to molecular weight or vice versa. Simply choose which state you expect the protein to be in and update the values accordingly.
Much like disposable cuvettes for DLS, we recommend that each chip is used just once. While the re-use of chips is not restricted, we can only vouch for the result obtained with the first use of each chip. This is because re-use increases the risk of introducing air into the system, as well as potential issues associated with protein adhesion and multiple actuations of the on-chip valve.
One cartridge contains enough reagents to run approximately 96 measurements which is equivilent to 4 chip boxes. The percentage remaining is indicated in the top right of the instrument display. Changing the cartridge takes a few minutes and does not require any special tools.
The exact protocol may vary depending on the software version of your instrument — check the user manual for full instructions on how to perform the change.
Reagent cartridges are disposable and should not be re-filled.
MDS exploits the unique properties of flow in microfluidic channels — specifically laminar flow, where streams can flow alongside one another with no convective mixing.
Watch the animation explaining MDS and the Fluidity One-W Serum below.
A selection of key scientific references and related content related to COVID-19, Coronaviruses, and SARS-CoV-2.
It is likely that changes in hydrodynamic radius (Rh) resulting from phosphorylation state changes would be too small to detect. However, if phosphorylation triggers a state change, such as dimerization, then this may be observed by monitoring Rh.
The hydrodynamic radius (Rh) determined by our Fluidity instruments is the Stokes radius, or the size of a hard spherical particle that diffuses at the same rate as the protein(s) detected.
For proteins, this is largely determined by the molecular weight, but shape also plays a role, with compact, well-folded proteins diffusing faster than extended, poorly folded proteins, and thus having a smaller hydrodynamic radius. The size determined by gel permeation or size exclusion chromatography is also the hydrodynamic radius.
Click here for details on how to convert hydrodynamic radius of proteins to molecular weight
The correlation of hydrodynamic radius with protein molecular weight (Mw) is dependent on protein conformation. If proteins are globular, a strong correlation between Rh and Mw is observed. If proteins are unstructured or elongated the measured Rh will be larger than that of a globular protein of identical Mw. Consequently, comparison to a calibration curve of globular proteins will yield a good approximation of Mw for a globular protein of unknown Mw. This principle is used to calibrate SEC columns using samples of known molecular weights as the calibration standards.
To learn how to convert between hydrodynamic radius and molecular weight visit our FAQ on the topic here, we also have a Hydrodynamic Radius Converter to help convert between molecular weight and hydrodynamic radius.
1. Insert USB stick into the port on the front of the unit next to the touchscreen
2. Log in to the user account where the data you wish to download is saved (If you are an admin you can download all data on the instrument)
3. Open the side bar menu by touching the three stripe icon in the top left corner, then select “View results”
4. Long press on the first result you want to download, and tick boxes will appear next to all of the results listed
5. Touch each tick box to select the results required, or alternatively touch “Select all” from the options bar displayed
6. Once all the required results are ticked touch “Export files”. The “Export options” dialog box will appear—by default results are saved as .json, but you can select here to also export as .csv or Microsoft Excel spreadsheets. Admin accounts only will also see the option “Also delete files after export”
7. A file browser dialog will appear, navigate to the folder you want to export results into and touch “Select”.
8. Once the chosen file formats are selected touch “Export files” to perform the export. A message saying “Files successfully exported” will appear when the export is finished.
Results will remain saved on the instrument after they have been downloaded. To delete results after download select view results from the side bar menu, and touch the small graph icon next to the result, and select “Delete result” from the options.
No. The flow rate is changed when the protein size is selected as you begin a measurement, but the auxiliary fluid is supplied with the reagent cartridge and remains the same.
If you would like to discuss custom reagent cartridges that include your auxiliary fluid of choice please contact us.
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