Evaluation of a New Single-Use UV Sensor for Protein A Capture

As the adoption of single-use systems continues to expand beyond bags and tubing to complete process steps, a full range of sensing technologies will be needed to complement the resulting varied single-use applications. Single-use sensors must meet or exceed the performance of traditional sensing technologies in areas such as accuracy, response time, ease of use, control system integration, process compatibility, regulatory requirements, and cost. Single-use flow-through process sensors are currently available for pressure, temperature, flow, and conductivity. Here, we report results from a comparative study of a new single-use flow-through UV sensor against traditional UV detection equipment in a protein A capture chromatography application.

Elements of UV Detection
Several types of chromatography steps are routinely used to separate components during downstream processing in biopharmaceutical manufacture. Protein A affinity chromatography with UV spectroscopic detection is one of the most important types, partly because of its strong affinity for antibodies.

PRODUCT FOCUS: ANTIBODIES

PROCESS FOCUS: DOWNSTREAM

WHO SHOULD READ: ANALYSTS, PROCESS MANAGERS

KEYWORDS: PURIFICATION, DISPOSABLES, DATA ACQUISITION, IN-LINE MONITORING

LEVEL: INTRODUCTION
UV detection is particularly useful in bioprocess applications because biomolecules can absorb specific wavelengths of light between 200 and 400 nm. In particular, proteins usually show strong absorbance around 280 nm because of the presence of aromatic amino acids in their structures.

According to the well-known Beer’s Law, the amount of light absorbed is directly proportional to the concentration of an absorbing molecule in the path length of the light. The path length through which the light travels is also important. For long path lengths, more of a light-absorbing molecule is present in the light’s path. A detector measures the ratio of incoming (incident) light energy to the energy that has passed through a sample as a measure of the absorbance of a material. A simple UV chromatogram compares time withabsorbance in a flow-through system as molecules in a solvent move through a UV light beam.

Table 1:Comparison of system design specifications 
In-line monitoring of UV absorbance during bioprocess chromatography is commonly applied for
• determination of when to start and end collection of a product-related peak
• confirmation of proper process execution during batch record review (by comparing a chromatogram with a reference chromatogram)
• identification and characterization of potential process disturbances during process investigations
• batch-to-batch process trending.
Often, accuracy at low-absorbance values


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