The American Chiropractic Board of Sports Physicians Position Statement on Pre-Participation Examinations: An Expert Consensus.

OBJECTIVE: The purpose of this paper is to present a position statement of best practices for the provision of a safe and high-quality pre-participation examination (PPE) and to provide recommendations on education requirements for doctors of chiropractic providing the PPE. METHODS: In 2014, the American Chiropractic Board of Sports Physicians (ACBSP) Board of Directors identified a need to review and update the ACBSP position statements and practice guidelines in order to be current with evolving best practices. Twelve ACBSP certificants, 10 Diplomates of the ACBSP, and 2 Certified Chiropractic Sports Physicians, met in April 2015 to author a pre-participation position statement using an expert consensus process. Panel members excluded anyone with commercial conflicts of interest and included individuals with expertise in clinical sports medicine and the performance of PPEs. A literature review was performed and circulated in advance for use by the panel in addressing the topic. The position statement was written through a consensus process and accepted by the ACBSP Board of Directors in May of 2015. RESULTS: The ACBSP Position Statement on Pre-participation Examinations identifies the qualifications and best practices for doctors of chiropractic to perform a PPE. CONCLUSION: This position statement states that doctors of chiropractic with post graduate education and current Diplomates of the ACBSP or Certified Chiropractic Sports Physicians certification have the prerequisite education and qualifying skills to perform PPEs.

Sports chiropractic management of concussions using the Sport Concussion Assessment Tool 2 symptom scoring, serial examinations, and graded return to play protocol: a retrospective case series.

OBJECTIVE: The purpose of this case series is to report how the symptom section of the Sport Concussion Assessment Tool 2 (SCAT2) was used to manage athletes with concussions in a high school training room setting and to address the need for SCAT2 baseline measurements. CLINICAL FEATURES: During a 4-month period, 3 doctors of chiropractic with certification from the American Chiropractic Board of Sports Physicians managed 15 high school athletes with concussions in a multidisciplinary setting. Fourteen athletes were male American football players, and one was a female volleyball player. INTERVENTION AND OUTCOME: Of the 15 athletes, 3 athletes had baseline SCAT2 documentation. Athletes were evaluated and returned to play with a graded return to play protocol using the SCAT2 symptoms and serial physical examinations. Once participants were asymptomatic, they began a graded return to play process. A total of 47 SCAT2 tests were performed on the 15 athletes, averaging 3.13 SCAT2 evaluations per patient. Of the 15 athletes evaluated, 6 were managed and cleared for return to play; 2 of the athletes sustained concussions in the last week of the season, thus ending their season; and 3 athletes were cleared by medical doctors. None of the athletes under care reported an adverse event. CONCLUSION: The utilization of the SCAT2 with serial physical examinations provided objective measures for athlete's injuries, allowing the practitioners to evaluate concussions. More efforts are needed to collect baseline SCAT2 to compare these scores with subsequent SCAT2 scores following athletic injuries.

Improved validation of peptide MS/MS assignments using spectral intensity prediction.

A major limitation in identifying peptides from complex mixtures by shotgun proteomics is the ability of search programs to accurately assign peptide sequences using mass spectrometric fragmentation spectra (MS/MS spectra). Manual analysis is used to assess borderline identifications; however, it is error-prone and time-consuming, and criteria for acceptance or rejection are not well defined. Here we report a Manual Analysis Emulator (MAE) program that evaluates results from search programs by implementing two commonly used criteria: 1) consistency of fragment ion intensities with predicted gas phase chemistry and 2) whether a high proportion of the ion intensity (proportion of ion current (PIC)) in the MS/MS spectra can be derived from the peptide sequence. To evaluate chemical plausibility, MAE utilizes similarity (Sim) scoring against theoretical spectra simulated by MassAnalyzer software (Zhang, Z. (2004) Prediction of low-energy collision-induced dissociation spectra of peptides. Anal. Chem. 76, 3908-3922) using known gas phase chemical mechanisms. The results show that Sim scores provide significantly greater discrimination between correct and incorrect search results than achieved by Sequest XCorr scoring or Mascot Mowse scoring, allowing reliable automated validation of borderline cases. To evaluate PIC, MAE simplifies the DTA text files summarizing the MS/MS spectra and applies heuristic rules to classify the fragment ions. MAE output also provides data mining functions, which are illustrated by using PIC to identify spectral chimeras, where two or more peptide ions were sequenced together, as well as cases where fragmentation chemistry is not well predicted.

Analysis of membrane proteins from human chronic myelogenous leukemia cells: comparison of extraction methods for multidimensional LC-MS/MS.

An important strategy for "shotgun proteomics" profiling involves solution proteolysis of proteins, followed by peptide separation using multidimensional liquid chromatography and automated sequencing by mass spectrometry (LC-MS/MS). Several protocols for extracting and handling membrane proteins for shotgun proteomics experiments have been reported, but few direct comparisons of different protocols have been reported. We compare four methods for preparing membrane proteins from human cells, using acid labile surfactants (ALS), urea, and mixed organic-aqueous solvents. These methods were compared with respect to their efficiency of protein solubilization and proteolysis, peptide and protein recovery, membrane protein enrichment, and peptide coverage of transmembrane proteins. Overall, approximately 50-60% of proteins recovered were membrane-associated, identified from Gene Ontology annotations and transmembrane prediction software. Samples extracted with ALS, extracted with urea followed by dilution, or extracted with urea followed by desalting yielded comparable peptide recoveries and sequence coverage of transmembrane proteins. In contrast, suboptimal proteolysis was observed with organic solvent. Urea extraction followed by desalting may be a particularly useful approach, as it is less costly than ALS and yields satisfactory protein denaturation and proteolysis under conditions that minimize reactivity with urea-derived cyanate. Spectral counting was used to compare datasets of proteins from membrane samples with those of soluble proteins from K562 cells, and to estimate fold differences in protein abundances. Proteins most highly abundant in the membrane samples showed enrichment of integral membrane protein identifications, consistent with their isolation by differential centrifugation.

Comparison of label-free methods for quantifying human proteins by shotgun proteomics.

Measurements of mass spectral peak intensities and spectral counts are promising methods for quantifying protein abundance changes in shotgun proteomic analyses. We describe Serac, software developed to evaluate the ability of each method to quantify relative changes in protein abundance. Dynamic range and linearity using a three-dimensional ion trap were tested using standard proteins spiked into a complex sample. Linearity and good agreement between observed versus expected protein ratios were obtained after normalization and background subtraction of peak area intensity measurements and correction of spectral counts to eliminate discontinuity in ratio estimates. Peak intensity values useful for protein quantitation ranged from 10(7) to 10(11) counts with no obvious saturation effect, and proteins in replicate samples showed variations of less than 2-fold within the 95% range (+/-2sigma) when >or=3 peptides/protein were shared between samples. Protein ratios were determined with high confidence from spectral counts when maximum spectral counts were >or=4 spectra/protein, and replicates showed equivalent measurements well within 95% confidence limits. In further tests, complex samples were separated by gel exclusion chromatography, quantifying changes in protein abundance between different fractions. Linear behavior of peak area intensity measurements was obtained for peptides from proteins in different fractions. Protein ratios determined by spectral counting agreed well with those determined from peak area intensity measurements, and both agreed with independent measurements based on gel staining intensities. Overall spectral counting proved to be a more sensitive method for detecting proteins that undergo changes in abundance, whereas peak area intensity measurements yielded more accurate estimates of protein ratios. Finally these methods were used to analyze differential changes in protein expression in human erythroleukemia K562 cells stimulated under conditions that promote cell differentiation by mitogen-activated protein kinase pathway activation. Protein changes identified with p<0.1 showed good correlations with parallel measurements of changes in mRNA expression.

Improving reproducibility and sensitivity in identifying human proteins by shotgun proteomics.

Identifying proteins in cell extracts by shotgun proteomics involves digesting the proteins, sequencing the resulting peptides by data-dependent mass spectrometry (MS/MS), and searching protein databases to identify the proteins from which the peptides are derived. Manual analysis and direct spectral comparison reveal that scores from two commonly used search programs (Sequest and Mascot) validate less than half of potentially identifiable MS/MS spectra (class positive) from shotgun analyses of the human erythroleukemia K562 cell line. Here we demonstrate increased sensitivity and accuracy using a focused search strategy along with a peptide sequence validation script that does not rely exclusively on XCorr or Mowse scores generated by Sequest or Mascot, but uses consensus between the search programs, along with chemical properties and scores describing the nature of the fragmentation spectrum (ion score and RSP). The approach yielded 4.2% false positive and 8% false negative frequencies in peptide assignments. The protein profile is then assembled from peptide assignments using a novel peptide-centric protein nomenclature that more accurately reports protein variants that contain identical peptide sequences. An Isoform Resolver algorithm ensures that the protein count is not inflated by variants in the protein database, eliminating approximately 25% of redundant proteins. Analysis of soluble proteins from a human K562 cells identified 5130 unique proteins, with approximately 100 false positive protein assignments.