Hi everyone, and thanks in advance for reading!
I want to determine oxidative stress human serum/plasma using protein damage; I´ve seen there are a lot of kits and methods, I want to try an ELISA for this, but as I read more and more papers, the methods described are many times vague and unclear.
Please, If someone is doing this in his Lab or have experience on this issue, can you please reach me a protocol for sample preparation and protein carbonylation detection?
It will be very useful as I see there are a lot of steps and I will save a lot of time (and money) on starting it up making my own mess...
Thank you all!
Dear Oscar, I can offer to you the method, which is favourably differing from the protein carbonilation detection on simplicity and speed. The preparation of a blood plasma sample before measurement of the degree of oxidative damage to proteins includes gel-chromatographic separation of low-molecular antioxidants and proceeds about 10 minutes. The measurement themselves by means of a temperature-controlled chemiluminometer (for example, minilum® L100) lasts about two minutes. The description of the measuring principle is given in the attachments „Patent US ARAP.pdf“ and „SFRR 1999-Summary.pdf“. The relevant publications you can find here:
https://www.researchgate.net/publication/12875169_Photochemiluminescent_detection_of_antiradical_activity_VI_Antioxidant_characteristics_of_human_blood_plasma_low_density_lipoprotein_serum_albumin_and_amino_acids_during_in_vitro_oxidation
https://www.researchgate.net/publication/296257917_Anti-radical_properties_of_oxidatively_modified_nucleotides_and_DNA_measured_in_a_chemiluminescence_detection_system_with_free_radical_azo-initiator
https://www.researchgate.net/publication/7627343_Photochemiluminescent_detection_of_antiradical_activity_VII_Comparison_with_a_modified_method_of_thermo-initiated_free_radical_generation_with_chemiluminescent_detection
https://www.researchgate.net/publication/263148204_Antioxidative_system_of_the_organism_and_thermo-initiated_chemiluminescence_method_for_quantitative_evaluation_of_its_state
https://www.researchgate.net/publication/315552642_Antioxidative_homeostasis_its_evaluation_by_means_of_chemiluminescent_methods
If you have further questions, do not hesitate to contact me: [email protected].
Best regards, I. Popov.
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