The concept of a lab-on-a-molecule, which was proposed just short of two decades ago, has captured the imagination of scientists. From originally being proposed as an AND logic gate driven by three chemical inputs as a direct way of detecting congregations of chemical species, the definition of what constitutes a lab-on-a-molecule has broadened over the years. In this review, molecules that can detect multiple analytes by fluorescence, among other techniques, are reviewed and discussed, in the context of molecular logic and multi-analyte sensing. The review highlights challenges and suggestions for moving the frontiers of research in this field to the next dimension.

A strategy is described for continuous monitoring of multiple latent tuberculosis infection (LTBI) biomarkers, specifically of interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α) and interleukin-2 (IL-2). Silver nanoparticles acting as mass signal amplifiers were linked to respective antibodies to form mass nanoprobes for increasing the mass loaded on the surface of the quartz crystal microbalance (QCM). This results in enhanced sensitivity. The mass nanoprobes can be oxidatively dissolved by hydrogen peroxide that avoided the steric hindrance caused by the scale effect of mass nanoprobes. This offers the option of signal recovery monitoring. By using this method, IFN-γ, TNF-α and IL-2 can be monitored serially. The frequency shifts caused by TNF-α, IFN-γ and IL-2, respectively, are reversible. Hence, the biomarkers can be continuously quantified. Compared to multichannel QCM sensing, the new method avoids acoustic interference and has a simplified instrumental setup. The assay is simple, accurate, sensitive, and inexpensive. Graphical abstract Silver nanoparticles as the mass signal amplifiers were linked with the antibodies to form mass nanoprobes for enhancing the monitoring sensitivity. With the introduction of H2O2 to dissolve the mass nanoprobes attached on sensing interface, a signal recovery QCM strategy is established for real-time and continuous monitoring of three LTBI biomarkers.

BACKGROUND: Nanoscaled aptamers (Aps), as short single-stranded DNA or RNA oligonucleotides, are able to bind to their specific targets with high affinity, upon which they are considered as powerful diagnostic and analytical sensing tools (the so-called \"aptasensors\"). Aptamers are selected from a random pool of oligonucleotides through a procedure known as \"systematic evolution of ligands by exponential enrichment\".
METHODS: In this work, the most recent studies in the field of aptasensors are reviewed and discussed with a main focus on the potential of aptasensors for the multianalyte detection(s).
RESULTS: Due to the specific folding capability of aptamers in the presence of analyte, aptasensors have substantially successfully been exploited for the detection of a wide range of small and large molecules (e.g., drugs and their metabolites, toxins, and associated biomarkers in various diseases) at very low concentrations in the biological fluids/samples even in presence of interfering species.
CONCLUSIONS: Biological samples are generally considered as complexes in the real biological media. Hence, the development of aptasensors with capability to determine various targets simultaneously within a biological matrix seems to be our main challenge. To this end, integration of various key scientific dominions such as bioengineering and systems biology with biomedical researches are inevitable.

A sensitive and quantitative fluorescent multi-component immuno-chromatographic sensor was developed for detection of three β-agonizts: clenbuterol, ractopamine and salbuterol. A competitive immune strategy between antibody conjugated fluorescent beads and β-agonist or their antigens was employed. Each monoclonal antibody specifically recognizes it is corresponding β-agonist in the conjugating zone. The unreacted antibodies were captured by β-agonist antigens immobilized at three test lines in nitrocellulose membrane reaction zone. This enables simultaneous detection of 3 β-agonizts in one single test without any further sample preparation. The test results can be obtained within 10 min. Limit of detections for clenbuterol, ractopamine and salbuterol were 0.10 ng/mL, 0.10 ng/mL and 0.09 ng/mL, respectively. Recoveries ranged from 70.0% to 100.5% and relative standard deviations were below 15%. The assay was evaluated using spiked and real samples and the results were compared with LC-MS/MS. The developed novel assay method provides a low cost, sensitive and rapid approach for on site detection of β-agonizts.

Mycotoxins are highly toxic contaminants and have induced health threat to human and animals. Aflatoxin B1 (AFB1), ochratoxin A (OTA) and zearalenone (ZEA) commonly occur in food and feed. A multi-component immunochromatographic assay (ICA) was developed for rapid and simultaneous determination of these three mycotoxins in agro-food. The strategy was performed based on the competitive immunoreactions between antibody-colloidal gold nanoparticle conjugate probes and mycotoxins or mycotixin antigens. Each monoclonal antibody specially recognize its corresponding mycotoxin and antigen, and there was no cross reactivity in the assay. Three mycotixin antigens were immobilized as three test lines in the nitrocellulose membrane reaction zone, which enable the simultaneous detection in one single test. The visible ICA results were obtained in 20 min. The visual detection limits of this strip test for the AFB1, OTA and ZEA were 0.25 ng/mL, 0.5 ng/mL and 1 ng/mL, respectively. The assay was evaluated using spiked and naturally contaminated peanuts, maize and rice samples. The results were in accordance with those obtained using enzyme-linked immunosorbent assay. In summary, this developed ICA could provide an effective and rapid approach for onsite detection of multi-mycotoxin in agro-food samples without any expensive instrument.