Technical Tips

We all like to keep as much biosensor activity during operational use as possible. One of the main ways that sensors lose sensitivity is that proteins and other substances foul the surfaces of the biosensor thus clogging the porous matrix and passivating the Pt surface and reducing electrochemical detection. The user can do two things to reduce this. Firstly dipping the biosensor tip into a 50% solution of PEG 20k (polyethyleneglycol mean molecular weight of 20,000) before use for a few minutes before washing off the excess PEG will help to reduce protein fouling. Secondly, cycling the sensor from -500 mV to +500mV and back again will help to regenerate the Pt surface of the electrode and restore electrochemical sensitivity. This procedure should be perfomed with the sensors out of contact with tissue.
Do you use sarissaprobes™ Ado, Ino or Hyp? If so, rather than using the standard phosphate buffer to store them in after rehydration, you may want to use a Tris buffer that gives better preservation of activity in wet storage. Composition of Tris buffer: 100mM NaCl, 1mM CaCl2, 1mM MgCl2, 20mM glycerol and 50 mM Tris, pH 7.4. (To make 1l of this buffer dissolve 6.777g Trizma hydrochloride, 0.822g Trizma base, 1ml CaCl2 (1M stock), 1ml MgCl2 (1M stock), 1.84g glycerol and 5.85g NaCl in a total volume of 1l.)

If you have difficulties making recordings with our probes, you may wish to check whether your recording system is set up correctly. There are two ways to check the instrumentation, one is to connect a high value resistor (e.g. 100 MOhms) across the terminals of your potentiostat and check whether the current recorded by the potentiostat at a particular voltage is in agreement with Ohm’s law. A second method is to apply H2O2, for example at 10 μM. All our probes (even the Null sensors) are sensitive to H2O2, therefore if your recording apparatus is functioning correctly you should see a large and rapid current response to H2O2. If you don’t see this, check that all connections to the potentiostat have been made correctly, the integrity of your leads and the bath for the Ag/AgCl reference electrode.

Preventing protein fouling
We all like to keep as much biosensor activity during operational use as possible. One of the main ways that sensors lose sensitivity is that proteins and other substances foul the surfaces of the biosensor thus clogging the porous matrix and passivating the Pt surface and reducing electrochemical detection. The user can do two things to reduce this. Firstly dipping the biosensor tip into a 50% solution of PEG 20k (polyethyleneglycol mean molecular weight of 20,000) before use for a few minutes before washing off the excess PEG will help to reduce protein fouling. Secondly, cycling the sensor from -500 mV to +500mV and back again will help to regenerate the Pt surface of the electrode and restore electrochemical sensitivity. This procedure should be perfomed with the sensors out of contact with tissue.

Storing ADO/INO/HYP sensors
Do you use sarissaprobes™ Ado, Ino or Hyp? If so, rather than using the standard phosphate buffer to store them in after rehydration, you may want to use a Tris buffer that gives better preservation of activity in wet storage. Composition of Tris buffer: 100mM NaCl, 1mM CaCl2, 1mM MgCl2, 20mM glycerol and 50 mM Tris, pH 7.4. (To make 1l of this buffer dissolve 6.777g Trizma hydrochloride, 0.822g Trizma base, 1ml CaCl2 (1M stock), 1ml MgCl2 (1M stock), 1.84g glycerol and 5.85g NaCl in a total volume of 1l.)

Checking your recording system
If you have difficulties making recordings with our probes, you may wish to check whether your recording system is set up correctly. There are two ways to check the instrumentation, one is to connect a high value resistor (e.g. 100 MOhms) across the terminals of your potentiostat and check whether the current recorded by the potentiostat at a particular voltage is in agreement with Ohm’s law. A second method is to apply H2O2, for example at 10 μM. All our probes (even the Null sensors) are sensitive to H2O2, therefore if your recording apparatus is functioning correctly you should see a large and rapid current response to H2O2. If you don’t see this, check that all connections to the potentiostat have been made correctly, the integrity of your leads and the bath for the Ag/AgCl reference electrode.

Using screened sensors effectively

The new internal screening layer cuts out the majority of interfering signals and makes the sarissaprobe® biosensors much more selective compared to the earlier versions. Nevertheless it is essential that the experimenter still performs controls (with appropriately chosen reference sensors) to establish the origin of the signal recorded by the biosensors and to eliminate the possiblity that it derives from some electroactive interferent. Note that the ATP-Null sensor (SA1002) which does not possess any enzymes can be used as a reference sensor for all sarissaprobe® sensors. More specific reference sensors for adenosine and acetylcholine measurements are sarissaprobe®-INO (SA1004) and sarissaprobe®-ACH-Null (SA1006) respecitvely.

The internal screening layer can become less effective during operational use and prolonged storage (>2 months dry). We strongly recommend testing the integrity of this layer with an electroactive substance such as 5-HT (e.g. at 10 µM). This is especially important if you wish to obtain differential recordings between sensors -both need to exhibit the same selectivity. If the screening has weakened, even although sensitivity to the analyte of interest is good, it is possible repair damage to the internal screening layer and restore selectivity by means of the following process:

Rescreening the sensors

For expert users only

This procedure should only be performed by users who have attained expertise in the use and handling of the sensors. Note that 1,3 diaminobenzene is harmful; be sure to follow the correct safety precautions for its handling and disposal. Do not use the same Ag/AgCl reference for rescreening and physiological measurements.

Improving operational stability of sensors

When used in tissue biosensors can lose sensitivity gradually, leading to limits on the operational stability and life of sensors. This is due to several factors, but adsorption of proteins onto the biosensor surface is an important contributor. The adsorbed proteins impede analyte access into the biosensor matrix and hence reduce sensitivity. This protein adsorption can be significantly reduced by an external coating of polyethyleneglycol (PEG). We recommend that the user dips the rehydrated biosensors into a 50% solution of PEG (20k molecular weight) for 10-15 mins before use with tissue. Excess PEG should then be rinsed off with pure water, and the sensors used in the normal way. This will protect the biosensors for several hours. If need be this procedure can be repeated each time the sensor is reused.

Specification