Analysis of Samples taken from the Cochlear Apex
The methods most commonly used to collect perilymph samples from the cochlea are either to perforate
the basal turn and take a sample or to insert a sampling pipette through the round window membrane in the basal turn of scala tympani and aspirate the sample.
In both cases the sample obtained is highly contaminated with cerebrospinal fluid (CSF). It is estimated that a 1 uL sample from a guinea pig
contains 30% CSF, a 2 uL sample contains 50% CSF and a 10 uL sample contains 85% CSF. Analysis of such samples cannot accurately reflect perilymph composition.
We have recently developed a better method that takes the perilymph sample from the cochlear apex. The goal of this method is to collect ALL the fluid that emerges from the cochlea when it is perforated,
thus ensuring that all the perilymph is collected. The basic elements of the method are shown below.
1) Remove mucosa from the apex and allow bone to dry
2) Apply thin cyanoacrylate adhesive to the apex and spread thinly.
3) Construct a silicone cup over the apex with only a thin (transparent) film of silicone over the site to be perforated.
Multiple (3-5) applications of WPI Kwik-Cast, 2-part silicone are required.
4) Make a small (50 - 100 ?m) fenestra at the apex with a Storz N170580, 1/3mm 30 degree oval window pick.
Fluid will immediately start to form a “bead” on the silicone.
5) Collect the fluid as it emerges into blunt tipped calibrated capillary (e.g. VWR # 53432-728 10 mL micropipette)
held by hand against the apex. Sample accumulates at 0.5 – 1 mL/min.
Fluid can be collected as a single large (10 uL) sample. Details of the large-sampling method and the interpretation of sample
concentrations are presently being published ( Salt, A.N., Hale, S.A., Plontke, S.K.R. Perilymph sampling from the cochlear apex: A reliable method to obtain higher purity perilymph samples from scala tympani. Journal of Neuroscience Methods in press, October 2005).
In addition, if the sensitivity of the measurement system permits, it is possible to take multiple smaller samples from the apex. The details of this method are presently being published.
As detailed in the publication (in preparation) sequential samples from the apex can be interpreted with our automated spreadsheet
that uses curve-fitting to estimate the basal-apical concentration gradient of drug.
Current release: File date December 9, 2005
The spreadsheet uses a macro. When the spreadsheet is run, Excel may require confirmation that macros should be enabled.
The spreadsheet also requires "Solver" to be functional in Excel.
Solver is a curve-fitting package which is not installed unless specifically requested.
If "solver" is not one of the options on the Tools menu in Excel, then you need to do a re-install of Excel,
finding the option that permits Solver to be installed.
The spreadsheet was written in Microsoft Excel 2000.