How Do Optical Dissolved Oxygen Sensors Work?

Dissolved oxygen (DO) is one of the most critical elements to measure in cell culture and also one of the most often overlooked. This oversight is for a variety of reasons ranging from lack of knowledge to the intrusive nature of traditional DO probes. However, for most cell culture conditions, there is a simple solution found in optical DO sensors. Though the technology was invented in the late 1990s, it was not until very recently that it became affordable enough to be used at large scale and available for any biomanufacturing team. When it comes down to the minute details of laboratory cell culture there are really only two types of DO sensors, electrochemical and optical.

Electrochemical DO sensors are most often referred to as Clark electrodes after Leland Clark, who invented the technology in 1962. And the optical sensor, developed almost four decades later, is named as you might assume for its reliance on optics. 

How do optical dissolved oxygen sensors work? 

Optical DO sensors are made of two parts, a sensor spot and a fiber optic reader. The sensor spot is attached to the interior of a cell culture vessel and contains a fluorescent dye suspended in a hydrogel. The sensor spot is the only part of the system that comes in contact with cells or media. The external reader is connected to a computer or data hub and is responsible for sending and receiving optical signals to the sensor spot. Optical DO sensors measure the DO concentration of liquid media or air based on the quenching of luminescence in the presence of oxygen. Since oxygen affects both the intensity and the lifetime of the luminescence, either may be used to measure DO. 

There are three methods of measuring optical dissolved oxygen:

• Magnitude domain measures the peak of luminescence. 

• Lifetime domain measures the decay rate of luminescence. 

• Phase domain measures the phase difference of the entire signal. 
  

Optical DO sensor technology was developed from the principle that DO quenches the luminescence associated with chemical dyes in the sensor. The fluorescent dye exhibits an excitation maximum around 455nm, and an emission wavelength of approximately 613nm. The fiber optic reader emits a blue light that excites the fluorophore which emits light back at the higher wavelength. In the presence of oxygen, the fluorescence is quenched, and a higher level of oxygen leads to lower emission intensities. This quenching also leads to changes in decay time which is the property that most readers use to determine DO levels.

The reader measures DO by emitting the blue light that causes the sensing element to luminesce and then the computer translates the signal decay. Lifetime domain-based sensors have some advantages over magnitude domain sensors and are the more common type of optical DO sensor on the market. They have better long-term accuracy and stability and are not as susceptible to leaching, dye degradation, or signal drift. 

What are the benefits of optical DO sensors? 

No flow required

Because optical sensors do not consume oxygen there is no need for a steady replenishment of oxygen at the tip of the sensor. This allows for the use of optical sensors in static cell culture. 

Minimal maintenance

The optical sensor spot is the only part of the device that comes in contact with cells or media and they are designed to be single use, affordable, and disposable. Since there is no membrane to replace and no cathode to take care of, regular maintenance is measured in months or years rather than days or weeks. 

No need for calibration

High quality optical DO sensors come pre-calibrated and do not need to be adjusted before use. This limits any possible down-time during experiments. 

Short response time

Optical sensors have a response rate similar to optical probes but since there is no need for daily calibration, they have a greater lifetime of measurements. 

What are the limitations of optical DO sensors?  

Sensor spot sensitivity

Optical sensor spots are small, delicate, and must be handled with care. Exposure to high concentrations of alcohols or certain organic compounds can also cause damage. 

Entry cost

While the lifetime cost of optical DO sensors is lower than other types of probes because of replacement and maintenance, the initial cost is usually higher than electrochemical probes.

New technology

Despite having been discovered more than 20 years ago, optical sensor technology is still relatively new to the market and many users are not familiar with how they work.