Comparación de métodos de extracción de proteínas de cerebro y linfocitos de rata
Comparison of protein extraction methods from brain and lymphocytes of rat
DOI:
https://doi.org/10.54167/tch.v11i3.87Palabras clave:
proteómica, electroforesis bidimensional (2-DE), extracción de proteínas, cerebro, linfocitosResumen
En este trabajo se compararon tres técnicas de extracción de proteínas actualmente empleadas en proteómica, para determinar la más eficiente para realizar electroforesis bidimensional (2-DE) en tejido cerebral y linfocitos de sangre periférica de rata. Los métodos utilizados fueron el uso directo de solución de lisis, el método TCA/acetona-DTT y el método TCA/acetona-fenol. Una vez que se realizó la extracción, se separaron las proteínas por medio de electroforesis en geles de poliacrilamida en condiciones desnaturalizantes (SDS-PAGE) y 2-DE, con el objetivo de seleccionar cuál de ellos brindó un mayor rendimiento en la cantidad de proteínas totales, así como en el número de bandas bien definidas y manchas bien enfocadas en los geles 2-DE, tanto para cerebro como para linfocitos. Al comparar el perfil proteico, en cerebro se detectaron 13 ± 0; 15 ± 1 y 12 ± 1 bandas bien definidas mediante los métodos de TCA/ acetona-DTT, TCA/acetona-fenol y solución de lisis, respectivamente. En linfocitos, se encontraron 19 ± 1.20 ± 0 y 19 ± 1 bandas, respectivamente. Con respecto al proteoma, tanto en cerebro como en linfocitos se encontró mayor número de manchas proteicas consistentes y bien enfocadas con el método de TCA/acetona-DTT. Estos resultados mostraron que el mejor método de extracción de proteínas para su uso en la 2-DE correspondió al de TCA/acetona-DTT, siendo además más rápido y sencillo de realizar que el método de TCA/acetona-fenol.
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