Part:BBa_K1086001:Experience

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Applications of BBa_K1086001

We have constructed the composite RCNA+YFP to detect IMA (ischemia modified albumin) in the serum of patients with cardiac risk. These patients present more IMA than normal individuals.

IMA binds less to cobalt than normal albumin, so we expected that a serum containing more IMA would have more free cobalt than a “normal” serum. This excess of cobalt would be able to activate more RCNA promoter, which in turn would lead to expression of YFP, so we could be able to distinguish between a patient with cardiac risk from a normal patient by comparing the fluorescence generate by each serum.

To prove that our construct works, we made some tests with transformed E. coli XL1-Blue. We added different concentrations of cobaltous chloride to bacterial cultures and measured their fluorescence (excitation: 514nm; emission: 527 nm) and absorbance (600 nm) for a certain period. The results (Figure 1) show a peak after 3 hours, during exponential phase.

We have also added mice serum to bacteria containing the composite. These sera were from ischemic or non ischemic mice. We could see that for ischemic animals the generation of fluorescence was much higher than for non ischemic mice (Figure 2).

Conclusion:

Our results show that the composite RCNA+YFP generates fluorescence in the presence of cobalt. Furthermore, it can be used to distinguish between ischemic and non ischemic individuals.

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