Ultrasensitive graphene FET aptasensor for direct attomolar detection of glutamate in human clinical samples

Glutamate, the principal excitatory neurotransmitter in the brain, is crucial for cognition and memory, and its dysregulation is implicated in several neurological disorders, including Alzheimer’s Disease and epilepsy. However, precise and high-throughput quantification of glutamate in physiological samples remains challenging. Here, we report an ultrasensitive and highly specific glutamate aptamer-based biosensor, or aptasensor, developed through in silico design and microfabrication, followed by in vitro and clinical validation. The biosensor consists of arrays of graphene field-effect transistors functionalized with a novel DNA aptamer, NG-Apt-Glu, designed and characterized computationally and biochemically, revealing two putative glutamate binding sites. The aptasensor detects glutamate in artificial cerebrospinal fluid with a 1 aM detection limit, a wide linear range (1 aM–10 pM), and 24 mV/decade sensitivity, showing strong selectivity against GABA, glutamine, dopamine, and serotonin. To evaluate clinical applicability, glutamate levels were measured in cerebrospinal fluid from patients with Alzheimer’s Disease, showing a significant increase relative to controls and correlating with neurofilament light chain concentrations, a biomarker of neuronal death. These findings underscore glutamate’s involvement in Alzheimer’s pathophysiology and its potential as a biomarker for neurodegeneration. This ultrasensitive graphene-based aptasensor enables point-of-care monitoring, paving the way for early diagnosis and the development of novel therapeutic strategies.