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The production of insects on an industrial scale has attracted the attention of the research and agricultural industry as novel protein sources. To detect the presence of Gryllodes sigillatus (GS) in feed and food, a real-time PCR method based on the mitochondrial cytochrome b (CYB) gene is proposed by this study. Forty DNA samples of animal and plant origin were used to confirm the specificity of the qPCR system. The detection method’s performance was evaluated on different processed GS matrices including native GS (UnGS) and different commercial products: crunchy roasted samples (RoGS), insect meal mixtures (ACGS) and energetic snacks containing GS (GSS). Data on sequencing were aligned with the reference gene to confirm the PCR products. The regression curve (y = −3.394 x + 42.521; R2 = 0.994, d.f. 14) between Ct values and Log DNA concentrations of Gryllodes sigillatus resulted in an efficiency of 96.4%. The severity of the technological processing treatments and the matrix structure affected the intensity of the PCR signal with the same amount of insect DNA as observed by different y-intercepts of the three-regression lines for RoGS, ACGS, and GSS. The real-time PCR method resulted in robust and sensitive outcomes able to detect low amounts of GS DNA (5 g/100 g) in a complex matrix, making it suitable for detecting the presence or absence of labeled Gryllodes sigillatus material both in feed and food.