The composition of wheat straw leaf and stem fractions were characterized using traditional strong acid hydrolysis, and monoclonal antibodies using comprehensive microarray polymer profiling (CoMPP). These results are then related to high throughput lignocellulose pretreatment and saccharification screening data. Pure leaf fraction of wheat straw was the least recalcitrant compared to pure stem and easily digested by commercial cellulases after moderate hydrothermal pretreatment; 63% and 31% (w/w) of glucan, 88% and 61% of xylan were released from the leaf and stem fractions, respectively. By preparing samples of various leaf-to-stem (L/S) ratios, we found shifting conversion behavior as processing parameters were modified. Increasing the enzyme dosage, pretreatment temperature and pretreatment time all significantly improved conversion rates in samples with more than 50% leaf content, whereas less impact was observed on samples with less than 50% leaf content. Enzyme affinity, desorption and readsorption with leaf and stem fractions may affect the sugar yield in wheat straw saccharification. The data suggest that the L/S ratio is an important parameter when adjusting or optimizing conversion processes and additionally in feedstock breeding. Furthermore, this highlights the need for rapid techniques for determining L/S ratio in wheat straw harvests. The CoMPP data on specific carbohydrates and leaf pectin highlight carbohydrate epitopes that may be useful as markers in the development of novel screening techniques; especially pectin or arabinogalactan proteins related epitopes are promising.