Electrochemistry of hot electrons in fully aqueous solutions at tetrahedral amorphous carbon thin film electrodes is discussed. The generation of these highly reducing chemical species was confirmed by normal pulse voltammetry and several electrochemiluminescent systems. Electron transfer into pre‐existing solvent cavities was observed at approximately −2.65 V vs. Ag/AgCl (sat.). Electrogenerated hot electrons were utilized as chemiluminescent mediators in heterogeneous sandwich immunoassay of Serum Amyloid A. The calibration curve was linear over four orders of magnitude and the detection limit was 85 ng L−1 that demonstrates the efficiency of hot electron generation at this electrode material.