Microfluidic Measurement of Antibody−Antigen Binding Kinetics from Low-Abundance Samples and Single Cells
Bibliographic record
Abstract
We present a simple microfluidic fluorescence bead assay for accurately measuring antibody-antigen binding kinetics with a standard inverted fluorescent microscope. We measured association and dissociation rate constants from antibody-antigen interactions spanning nearly 4 orders of magnitude in equilibrium binding affinity (30 pM-100 nM). Two versions of this assay are presented, which allow for dissociation rate measurements either directly, by use of fluorescently labeled antigen, or indirectly, by use of unlabeled antigen. We also demonstrate simultaneous, multiplexed binding measurements of multiple antibody-antigen interactions using a combination of spectral separation and spatial localization. Complete antibody-antigen binding kinetics were measured for as little as 8 × 10⁴ antibody molecules (~132 zeptomoles) immobilized on a single bead and less than 2 × 10⁶ antibodies (~3 attomoles) loaded into the microfluidic device, a reduction in detection limit and sample consumption of 4 orders of magnitude when compared to surface plasmon resonance (SPR) spectroscopy and alternative measurement techniques. We show that the microfluidic bead assay, when combined with small volume compartmentalization, enables direct measurement of antigen binding kinetics of antibodies secreted from single hybridoma cells. We anticipate that this assay will be useful as a routine analytical tool for studying molecular interactions as well as for screening primary antibody-secreting plasma cells isolated from immunized animals.
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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 | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 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".