An Enhanced Mass Spectrometry Approach Reveals Human Embryonic Stem Cell Growth Factors in Culture
Bibliographic record
Abstract
The derivation and long-term maintenance of human embryonic stem cells (hESCs) has been established in culture formats that are both dependent and independent of support (feeder) cells. However, the factors responsible for preserving the viability of hESCs in a nascent state remain unknown. We describe a mass spectrometry-based method for probing the secretome of the hESC culture microenvironment to identify potential regulating protein factors that are in low abundance. Individual samples were analyzed several times, using successive mass (m/z) and retention time-directed exclusion, without sampling the same peptide ion twice. This iterative exclusion -mass spectrometry (IE-MS) approach more than doubled protein and peptide metrics in comparison to a simple repeat analysis method on the same instrument, even after extensive sample pre-fractionation. Furthermore, implementation of the IE-MS approach was shown to enhance the performance of an older quadrupole time of flight (Q-ToF) MS. The resulting number of identified peptides approached that of a parallel repeat analysis on a newer LTQ-Orbitrap MS. The combination of the results of both instruments proved to be superior to that achieved by a single instrument in the identification of additional proteins. Using the IE-MS strategy, combined with complementary gel- and solution-based fractionation methods, the hESC culture microenvironment was extensively probed. Over 10 to 12 times more extracellular proteins were observed compared with previously published surveys. The detection of previously undetectable growth factors, present at concentrations ranging from 10(-9) to 10(-11) g/ml, highlights the depth of our profiling. The IE-MS approach provides a simple and reliable technique that greatly enhances instrument performance by increasing the effective depth of MS-based proteomic profiling. This approach should be widely applicable to any LC-MS/MS instrument platform or biological system.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.001 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.001 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".