Question

Why do we need to create a calibration curve for the Spectrophotometer before we test our samples from the intestine?

Answer 1

Creating a calibration curve for the spectrophotometer is crucial for accurately estimating cell density based on turbidity, as it allows one to correlate turbidity readings with actual cell counts, using known standards for comparison.

Step-by-step explanation:

It is important to create a calibration curve for the spectrophotometer before testing samples to accurately estimate the cell density from turbidity measurements. The calibration curve is established by using known concentrations (with viable plate count data) to correlate optical density (turbidity) as a function of cell density.

Measuring the transmission of light through a control tube containing only broth helps to correct for any absorbance due to the medium itself rather than the cells. This is key for ensuring that any increase in absorbance measured by the spectrophotometer is due to bacterial cells present in the sample. Without this calibration step, the turbidity readings cannot be confidently associated with the actual number of cells.

The purpose of a calibration curve is to provide a reference that can be used to estimate cell counts for similar samples. This is essential as it allows for quick estimations of cell density in various samples without the need for time-consuming viable counts for each new measurement.

Answer 2

Creating a calibration curve for the spectrophotometer is crucial for accurate estimation of cell density from turbidity measurements. It establishes a baseline to differentiate between the turbidity of the medium itself and that caused by bacterial cells, allowing for accurate cell concentration assessments in biological samples.

Step-by-step explanation:

The creation of a calibration curve for the spectrophotometer is essential before testing samples from the intestine because it allows for the accurate estimation of cell density based on turbidity measurements. When using a spectrophotometer, transmission of light and increase in absorbance are measured. A calibration curve is generated by taking absorbance readings (which are related to cell density) from known standard solutions.

This process ensures that subsequent measurements of samples can be quickly and accurately associated with cell concentrations. It is particularly important in biological samples, such as those from the intestine, where cell density relates to the function and health of the tissue. Additionally, calibrating with a control tube containing only broth is critical because it establishes a baseline for light scattering that is due solely to the medium, excluding any effects from bacterial cells. The calibration curve thus allows researchers to differentiate between the inherent turbidity of the medium and the turbidity caused by the bacterial cells in the actual sample.