Isotyping immunoglobulins using accurate molecular mass

A method for detecting immunoglobulin heavy chains in a sample, the method comprising:

• a) providing a biological sample comprising immunoglobulins, paired immunoglobulin heavy and light chains, or mixtures thereof,
• b) immunopurifying the sample utilizing a single domain antibody fragment (SDAF) having affinity for an immunoglobulin, wherein said SDAF is selected from the group consisting of an anti-human IgG SDAF, an anti-human IgA SDAF, an anti-human IgM SDAF, an anti-human IgD SDAF, an anti-human IgE SDAF, an anti-human kappa SDAF, an anti-human lambda SDAF, and combinations thereof;
• c) subjecting the immunopurified sample to a decoupling step where light chain immunoglobulins are decoupled from the heavy chain immunoglobulins;
• wherein one or more of the immunoglobulin light chains or immunoglobulin heavy chains is derived from an M-protein;
• d) subjecting the immunopurified sample to a mass spectrometry technique to obtain a mass spectrum of the sample, said mass spectrum comprising one or more peaks corresponding to one or more intact immunoglobulin light chains in the sample;
• wherein said one or more peaks quantify the amount of the one or more intact immunoglobulin heavy chains in the sample,
• wherein the mass spectrometry technique is chosen from the group consisting of (i) liquid chromatography electrospray ionization coupled to mass analyzer (ii) a microflow liquid chromatography electrospray ionization coupled to a quadrupole time-of-flight mass spectrometry technique and (iii) a matrix assisted laser adsorption ionization-time of flight mass spectrometry technique;
• and
• e) determining one or more of (i) the identity of the M-protein, (ii) the quantity of the M-protein, (iii) the pairing of immunoglobulin heavy chains and immunoglobulin light chains of the M-protein, and (iv) the quantitative amount of one or more of the immunoglobulin heavy chains and M-protein in the sample.

A method for monitoring treatment of a disorder in a subject, said method comprising:

• a) providing a biological sample obtained from the subject, wherein the biological sample comprises immunoglobulins, paired immunoglobulin heavy and light chains, or mixtures thereof;
• b) immunopurifying the sample, wherein the immunopurifying comprises using an antibody selected from the group consisting of an anti-human IgG antibody, an anti-human IgA antibody, an anti-human IgM antibody, an anti-human IgD antibody, an anti-human IgE antibody, and combinations thereof;
• c) subjecting the immunopurified sample to a decoupling step wherein immunoglobulin light chains are decoupled from immunoglobulin heavy chains;
• and
• d) subjecting the decoupled sample to a mass spectrometry technique to obtain a mass spectrum of the sample, said mass spectrum comprising one or more peaks corresponding to one or more intact immunoglobulin heavy chains or one or more intact immunoglobulin light chains in the sample;
• wherein said one or more peaks quantify the amount of the one or more intact immunoglobulin heavy chains or one or more intact immunoglobulin light chains in the sample.