Interferon-γ is a proinflammatory cytokine, and its production is related with effective host defense against intracellular pathogens. Therefore, the level of interferon-γ is considered a good biomarker for intracellular infections. It is also useful for the assessment, treatment progression and follow-up of non-communicable diseases, including cancer and autoimmune disorders, among others. This work addresses the development of a novel interferon-γ release assay (IGRA) to evaluate the expression of interferon-γ transcripts as biomarker produced by isolated T cells, as a main advantage. The method sequentially combined three different types of magnetic separation, including the immunomagnetic separation of the T cells performed on antiCD3 modified magnetic particles, the retrotranscription and multiplex double-tagging PCR on polydT-modified magnetic particles and, finally, the electrochemical genosensing on streptavidin magnetic particles as a support. This approach is able to quantify the levels of cellular interferon-γ produced by as low as 150 T cells with outstanding analytical features. The detection of interferon-γ transcripts is performed from only 100 μL of whole blood which can be potentially obtained by fingerprick, demonstrating a further clear advantage to be considered as a promising strategy for the quantification of this important biomarker in several clinical applications.

干扰素-γ是促炎细胞因子，其产生与有效的宿主防御细胞内病原体有关。因此，γ-干扰素水平被认为是细胞内感染的良好生物标志物。它对于非传染性疾病（包括癌症和自身免疫性疾病）的评估，治疗进展和随访也很有用。这项工作解决了开发新型干扰素-γ释放测定法（IGRA）的主要优势，该测定法可评估作为分离的T细胞产生的生物标志物的干扰素-γ转录本的表达。该方法顺序地结合了三种不同类型的磁分离，包括对抗CD3修饰的磁性粒子进行的T细胞的免疫磁分离，在polydT修饰的磁性粒子上进行逆转录和多重双标签PCR，最后在链霉亲和素磁性粒子上进行电化学基因检测。这种方法能够定量分析低至150 T细胞产生的细胞干扰素γ的水平，并具有出色的分析功能。干扰素-γ转录物的检测仅从100μL的全血中进行，而全血可以通过手指刺法获得，这显示出进一步的明显优势，可以认为是在一些临床应用中对该重要生物标志物进行定量分析的有前途的策略。这种方法能够定量分析低至150 T细胞产生的细胞干扰素γ的水平，并具有出色的分析功能。干扰素-γ转录物的检测仅从100μL的全血中进行，而全血可以通过手指刺法获得，这显示出进一步的明显优势，可以认为是在一些临床应用中对该重要生物标志物进行定量分析的有前途的策略。这种方法能够定量分析低至150 T细胞产生的细胞干扰素γ的水平，并具有出色的分析功能。干扰素-γ转录物的检测仅从100μL的全血中进行，而全血可以通过手指刺法获得，这显示出进一步的明显优势，可以认为是在一些临床应用中对该重要生物标志物进行定量分析的有前途的策略。