Methods are disclosed to detect, characterize, measure, and quantify human and humanized antibodies, and their conjugates, that may be present in pre-clinical animal biological samples, or human biological samples, including plasma/serum and tissue samples.

Provided herein are IEF focusing methods for determining the number of drug molecules present in at least one antibody-drug conjugate (ADC) species subpopulation. In one embodiment, the method comprises performing free solution isoelectric focusing on a sample comprising at least one ADC species, to obtain a focused sample. The absorbance of the sample at two different wavelengths is then measured, for example, over a range of pI values. Absorbance values at the two different wavelengths are compared at at least one corresponding pI value, where the at least one corresponding pI value is the pI of the ADC subpopulation. The number of drug molecules in the at least one ADC species subpopulation is then determined based on the comparison. The methods provided herein can also be used to determine the number of specific binding pair members bound to its target specific binding pair member.

Provided are a kit for measuring a titer of a human antibody, a humanized antibody, or a human Fc-fusion protein in human plasma or serum, and a method for measuring a titer of a human Fc-containing protein in human plasma or serum using the same, and more specifically, a kit for measuring a titer of human Fc-containing protein in human plasma or serum, the kit including a diluent for sample, a diluent for conjugate, and a cleansing solution and being used in analysis through an indirect enzyme-linked immunosorbent assay (indirect ELISA), and a method for measuring a titer of human Fc-containing protein in human plasma or serum using the same.

The present invention provides a highly sensitive in vitro method for diagnosing injury to the central nervous system (CNS), such as a traumatic brain injury (TBI) or stroke. The method involves contacting a sample of blood from a subject suspected of suffering a CNS injury event with at least one antibody capable of detecting a proteolytic fragment of the biomarker Protein Kinase C, gamma isoform (PKCg or PKCγ), which fragment corresponds to a proteolytic fragment of PKCg formed in the excitotoxic environment resulting from neuronal damage. Also disclosed are novel anti-PKCg antibodies useful in the diagnostic methods of the invention to provide diagnosis of CNS injury with essentially 100% accuracy.

The present invention provides methods, devices, compositions (e.g., capture complexes), and kits useful for enhancing the detection of antibodies in a test sample. The methods, devices, and compositions utilize detectable Fc-binding molecules such as Protein A, Protein G, and/or an Fc-specific antibody to amplify the signal of a detected antibody in immunoassays, such as lateral flow assays.

Provided is a biomarker for diagnosing at-risk mental state (ARMS) that may include one or more selected from the group consisting of biopyrrin, cortisol, a KFLC or a fragment thereof, and a AFLC or a fragment thereof. Further provided is an ARMS diagnosis of a subject that may be performed quickly, easily, and accurately by measuring the amount of the biomarker for diagnosing ARMS in a biological sample.

A bivalent binding agent, capable of binding a polypeptide dimer, consisting of two monovalent binders linked to each other via a linker, the first monovalent binder binds an epitope of a first target polypeptide comprised in said dimer and the second monovalent binder binds to an epitope of a second target polypeptide comprised in said dimer. Each monovalent binder has a Kdiss in the range of 5×10.sup.−3/sec to 10.sup.−4/sec, and the bivalent binding agent has a Kdiss of 3×10.sup.−5/sec or less. Methods of making and using such bivalent binding agent in histological staining procedures are also disclosed.

This invention provides, and in certain specific but non-limiting aspects relates to: —assays that can be used to predict whether a given ISV will be subject to protein interference as described herein and/or give rise to an (aspecific) signal in such an assay (such as for example in an ADA immunoassay). Such predictive assays could for example be used to test whether a given ISV could have a tendency to give rise to such protein interference and/or such a signal; to select ISV's that are not or less prone to such protein interference or to giving such a signal; as an assay or test that can be used to test whether certain modification(s) to an ISV will (fully or partially) reduce its tendency to give rise to such interference or such a signal; and/or as an assay or test that can be used to guide modification or improvement of an ISV so as to reduce its tendency to give rise to such protein interference or signal; —methods for modifying and/or improving ISV's to as to remove or reduce their tendency to give rise to such protein interference or such a signal; —modifications that can be introduced into an ISV that remove or reduce its tendency to give rise to such protein interference or such a signal; —ISV's that have been specifically selected (for example, using the assay(s) described herein) to have no or low(er)/reduced tendency to give rise to such protein interference or such a signal; —modified and/or improved ISV's that have no or a low(er)/reduced tendency to give rise to such protein interference or such a signal.

Engineered multivalent and multispecific binding proteins, methods of making, and their uses in the prevention, diagnosis, and/or treatment of disease are provided.

The present inventors have successfully found a large number of Fc variants with remarkably increased binding activity against and/or binding selectivity for mouse FcγRII by introducing amino acid alteration(s) to the Fc region.