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High‐dimensional analyses of cancers can potentially be used to better define cancer subtypes, analyze the complex tumor microenvironment, and perform cancer cell pathway analyses for drug trials. Unfortunately, integrated systems that allow such analyses in serial fine needle aspirates within a day or at point‐of‐care currently do not exist. To achieve this, an integrated immunofluorescence single‐cell analyzer (i2SCAN) for deep profiling of directly harvested cells is developed. By combining a novel cellular imaging system, highly cyclable bioorthogonal FAST antibody panels, and integrated computational analysis, it is shown that same‐day analysis is possible in thousands of harvested cells. It is demonstrated that the i2SCAN approach allows comprehensive analysis of breast cancer samples obtained by fine needle aspiration or core tissues. The method is a rapid, robust, and low‐cost solution to high‐dimensional analysis of scant clinical specimens. [ABSTRACT FROM AUTHOR]

Rapid analysis of single and scant cell populations is essential in modern diagnostics, yet existing methods are often limited and slow. Herein, we describe an ultra‐fast, highly efficient cycling method for the analysis of single cells based on unique linkers for tetrazine (Tz)/trans‐cyclooctene (TCO)‐mediated quenching. Surprisingly, the quenching reaction rates were more than 3 orders of magnitude faster (t1/2 <1 s) than predicted. This allowed multi‐cycle staining and immune cell profiling within an hour, leveraging the accelerated kinetics to open new diagnostic possibilities for rapid cellular analyses. [ABSTRACT FROM AUTHOR]