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As one of the most representative trade commodities along the ancient Silk Road, silk artifacts have long been studied for provenance using art historical methodologies, yet have lacked scientifically rigorous analytical techniques grounded in the natural sciences. Radiogenic strontium (Sr) isotopes (87Sr/86Sr), known for their distinct geographical signatures, have emerged as a promising provenance tracing tool, with applications extending from geology into archaeology. Silkworm cocoons, the primary textile material for silk, exhibit high organic content (>95%) and low Sr concentrations (~1 μg/g). Conventional geological analysis methods typically require large sample sizes (entire or multiple cocoons or several hundred milligrams), making them unsuitable for minimally destructive analysis of precious silk artifacts. In this study, we developed a technique for high-precision Sr isotope analysis tailored to trace amounts of silk samples. By combining strong oxidant digestion (HNO3-H2O2) with Sr-specific resin purification, we achieved efficient organic matrix removal while maintaining high strontium recovery (>99%) and ultra-low procedural blanks (≤35 pg). Purified samples were analyzed using thermal ionization mass spectrometry (TIMS) with a “sandwich” loading technique and silicotungstic acid emitter, enhancing ionization efficiency to 10%. The method reliably measured as little as 0.5 ng of the NBS987 standard, yielding a mean 87Sr/86Sr ratio of 0.710251 ± 0.000033 (n = 32, 2SD), consistent with certified values. This approach reduces sample consumption to approximately 10 mg per analysis, representing a 1–2 order of magnitude improvement over conventional methods. This technique was then applied to cocoons of various silkworm breeds from China’s five major silk-producing regions. The results revealed that Sr isotopic signatures of silk are influenced by both geographic origin and silkworm variety, with regional factors playing a dominant role. This confirms the viability of Sr isotopes as silk provenance indicators. Our technical breakthroughs in Sr isotope analysis of trace silk samples provide a foundational analytical framework for minimally invasive provenance studies of silk artifacts. The BRIEF REPORT is available for this paper at http://www.ykcs.ac.cn/en/article/doi/10.15898/j.ykcs.202504060079.