Technical Abstract: Cotton production and usage is a global enterprise. Most of the cotton produced in the U.S. today is exported globally, primarily to China. Improved and fast response quality measurement systems are needed in the global marketplace to assess the quality of cotton. One key need is the development of new field and at-line quality measurements that will lead to the vertical integration of the cotton quality chain from the cotton field to final fabric. Recent advances in portable Near Infrared (NIR) instrumentation could result in improved, rapid, and precise quality tools for laboratory and breeder analyses. A program was implemented to determine the ability of portable NIR instrumentation to monitor critical cotton fiber properties of breeder samples in the laboratory, at-line, and in the field, with initial emphasis on micronaire. Micronaire is an indirect measurement of the cotton fiber’s maturity and fineness (diameter). Comparative evaluations demonstrated good NIR spectral and between-instrument agreement for both portable and bench-top units for micronaire, and distinct NIR spectral differences between samples with varying micronaire were observed. The NIR measurement was fast and easy to perform. Using a portable, hand held NIR analyzer, a comparative evaluation determined the optimal instrumental settings, operational conditions, and laboratory and field measurement protocols. Field sampling systems were developed, and the NIR measurements of seed cotton for micronaire were performed directly on the cotton boll in several cotton fields. Distinct micronaire differences were detected between cotton varieties and bolls with known differences.