Tumor acidosis is one of the hallmarks indicating the initiation and progression of various cancers. Here, we present a protocol for preparing a hyperpolarized (HP) ¹³C-bicarbonate tissue pH MRI imaging contrast agent to detect aggressive tumors. We describe the steps for the formulation and polarization of a precursor molecule ¹³C-glycerol carbonate (¹³C-GLC), the post-dissolution reaction, and converting HP ¹³C-GLC to an injectable HP ¹³C-bicarbonate solution. We then detail procedures for MRI data acquisition to generate tumor pH maps for assessing tumor aggressiveness. For complete details on the use and execution of this protocol, please refer to Mu et al.¹.

This study aimed to implement a multimodal ¹H/HP-¹³C imaging protocol to augment the serial monitoring of patients with glioma, while simultaneously pursuing methods for improving the robustness of HP-¹³C metabolic data. A total of 100 ¹H/HP [1-¹³C]-pyruvate MR examinations (104 HP-¹³C datasets) were acquired from 42 patients according to the comprehensive multimodal glioma imaging protocol. Serial data coverage, accuracy of frequency reference, and acquisition delay were evaluated using a mixed-effects model to account for multiple exams per patient. Serial atlas-based HP-¹³C MRI demonstrated consistency in volumetric coverage measured by inter-exam dice coefficients (0.977 ± 0.008, mean ± SD; four patients/11 exams). The atlas-derived prescription provided significantly improved data quality compared to manually prescribed acquisitions (n = 26/78; p = 0.04). The water-based method for referencing [1-¹³C]-pyruvate center frequency significantly reduced off-resonance excitation relative to the coil-embedded [¹³C]-urea phantom (4.1 ± 3.7 Hz vs. 9.9 ± 10.7 Hz; p = 0.0007). Significantly improved capture of tracer inflow was achieved with the 2-s versus 5-s HP-¹³C MRI acquisition delay (p = 0.007). This study demonstrated the implementation of a comprehensive multimodal ¹H/HP-¹³C MR protocol emphasizing the monitoring of steady-state/dynamic metabolism in patients with glioma.

The bacteriophage population is large, dynamic, ancient, and genetically diverse. Limited genomic information shows that phage genomes are mosaic, and the genetic architecture of phage populations remains ill-defined. To understand the population structure of phages infecting a single host strain, we isolated, sequenced, and compared 627 phages of Mycobacterium smegmatis. Their genetic diversity is considerable, and there are 28 distinct genomic types (clusters) with related nucleotide sequences. However, amino acid sequence comparisons show pervasive genomic mosaicism, and quantification of inter-cluster and intra-cluster relatedness reveals a continuum of genetic diversity, albeit with uneven representation of different phages. Furthermore, rarefaction analysis shows that the mycobacteriophage population is not closed, and there is a constant influx of genes from other sources. Phage isolation and analysis was performed by a large consortium of academic institutions, illustrating the substantial benefits of a disseminated, structured program involving large numbers of freshman undergraduates in scientific discovery.