FAQ - Basic Research

 Microdialysis Technique

Question:  What is recovery?
Answer:

The recovery of a particular substance is defined as the concentration in the dialysate expressed as percent of the concentration in the interstitial fluid.

 

Question: How does the length of the membrane affect the recovery?
Answer:

A longer membrane and a lower flow rate will give a higher recovery.

 

Question: How shall the perfusion fluid be composed?
Answer:

Ideally it should be as close as possible to the composition of the extracellular fluid. However, you may want to change the concentration of sodium, potassium or calcium in order to influence the membrane function in the region you are studying.
CMA can offer Perfusion Fluid T1 for peripheral tissue and Perfusion Fluid CNS for brain tissue.

 

Question: What is the pH of the CNS perfusion fluid?
Answer:

The CNS perfusion fluid is intentionally not buffered in order to allow it to take on the same pH as the brain’s interstitial fluid. The pH of a non-buffered solution varies between 5-8.

 

Question: How does the pH of the CNS perfusion fluid impact the tissue?
Answer:

Since our perfusion fluid is unbuffered it can hardly itself impact the pH in the tissue. The perfusate will take on the same pH as the surrounding tissue. But the buffered substances in the tissue can have different recovery over the membrane which indirectly can have a small effect on the pH of the tissue.

 

Question: If the cut-off of the probe membrane is 20,000 Daltons, why do I not have 100% recovery for molecules of 20,000 Daltons?
Answer:

The ability of molecules to pass the membrane decreases logarithmically with the increase in molecular weight. By experience we know that most substances with a molecular weight up to 5,000 Da can be dialyzed when using a 20,000 Da membrane. This is of course very dependent on the substance and the sensitivity of the analytical method.

 

 Probes

Question: Which length of the microdialysis probe membrane shall I use?
Answer:

A longer membrane gives a better recovery of the substances you are interested in but the choice is usually limited by the size of the structure you want to study.
CMA can offer a variety of probes with different membrane lengths from 1mm to 10 mm suitable for most experiments.

 

Question: Which membrane material is to be recommended?
Answer:

A membrane with low cut off purifies your sample by excluding large molecules. A membrane with high cut off recovers large substances such as peptides or smaller proteins. Note that some substances can also bind to the membrane material. To optimize your choice you should make an in vitro test with the substance you want to monitor.
CMA can offer membranes made of PAES, polyarylethersulfone, 20,000 Daltons, cuprophane, 6,000 Daltons and PES polyetersulfone, 100,000 Daltons cut-off.

 

Question: You have a number of different Microdialysis probes? How shall I choose the right one for my experiment?
Answer:

There are of course many aspects which determines the choice. This is a brief overview, for more information see our catalog or product information on CMA web pages.
A stiff probe is suitable for a stereotaxic experiment on the brain while a flexible probe may be suited for microdialysis in a peripheral organ such as adipose tissue, muscle, liver or kidney.
CMA 12 the optimized probe for CNS use, ideal for chronic implantation
CMA 11 a thin stiff probe for discrete brain regions
CMA 7 an extremely small stiff probe for CNS studies in smaller animals such as mice
CMA 20 a soft non-metallic probe for peripheral tissues and blood vessels
CMA 30 a linear probe suitable for skin and other peripheral tissues
CMA 31 a linear probe suitable for skin and other peripheral tissues with 55k Da cut-off

 

Question: What perfusion flow rate should be used?
Answer:

A high flow rate if you want to remove or introduce as many molecules as possible per time unit.
A low flow rate should be used when you want to obtain a more concentrated dialysate (high recovery). Note that a low flow rate gives smaller volume. Consider also the volume needed for the analysis.

 

Question: What time is needed to obtain steady state conditions?
Answer:

The introduction of a probe into the tissue will always cause damage and the recovery of function will take a certain time period. An hour is often used to reach “baseline conditions”.

 

Question: Are the probes reusable?
Answer:

The preclinical probes can be used repeatedly if rinsed and stored in deionized water between experiments. However CMA can only guarantee single usage.

 

Question: When do I need the guides?
Answer:

Guides are used when you shall perform Microdialysis in the brain of a freely moving animal.

 

Question: How do I handle FEP tubing and Tubing adapters?
Answer:

When using a FEP tubing you shall cut the tubing with a sharp scalpel or similar to be sure the ends are open. After use rinse it with de ionized water to wash out the salts.
Tubing Adapters can be swelled in 70% ethanol for easy connections with tubing and syringes. They will shrink back in air again and ensure tight fit and zero internal volumes.

 

Question: I get faulty flow. What have I done?
Answer:

Maybe you have not calibrated the pump to the syringe you are using.
The syringe can be leaking or you are not using the syringe recommended by CMA.

 

Question: Why do I get faulty volume of my microdialysis sample?
Answer:

The probe may be leaking, the tubing may be blocked, something may be wrong with the pump, or the tubing adapters may be leaking.

 

Instruments

Question: What is the difference between the CMA 400 pump and the CMA 402 pump?
Answer:

CMA 400 has a pulse free flow from 1 nL/min  - 1 mL/min, runs four syringes simultaneously. The pump is calibrated for various sizes of syringes. Apart from microdialysis experiments, it can be used for microinjections of preset volumes that can be repeated in intervals.
CMA 402 has  flow rates between 0,1µL - 20 µL/min, run by two syringes individually (1, 2.5 or 5 ml) . Start/stop and flow rate can be set independent for each syringe.
Both pumps can be used as both push and pull option.

 

Question: What is the difference between CMA 142 and CMA 470 Fraction Collectors?
Answer:

The CMA 142 collects fractions from 1 probe, 20 fractions or from 2 probes, 10 fractions. Fraction volumes from 1uL - 50 uL in open vials.
CMA 470 is a refrigerated fraction collector, can collect 64 fractions from 1 up to 4 probes simultaneously (16 fractions/probe = 64 total). Fraction volumes from 1 uL - 2000 uL. Both opened and seal vials can be used.

 

Question: When shall the CMA 150 be used?
Answer:

The CMA 150 keeps a stable temperature of the animal during an anesthetized microdialysis study. This ensures a better quality of your study.

 

Question: What is the use of a CMA 120 System for Freely Moving Animals?
Answer:

This system enables Microdialysis studies on conscious, small laboratory animals over a long period of time. It can be used in combination with any one of CMA’s preclinical instruments.