IDENTIFICATION OF PROTEINS IN LIPOFUSCIN USING ANTIBODIES WITHIN THE HUMAN PROTEIN ATLAS Evelina Sjöstedt 1 , Dijana Djureinovic 1, Jan Mulder 2 and Caroline Kampf 1 1. Department of Immunology, Genetics and Pathology, Human Protein Atlas, Science for Life Laboratory, Uppsala University, Uppsala, Sweden 2. Department of Neuroscience, Human Protein Atlas, Science for Life Laboratory, Karolinska Institute, Stockholm, Sweden Background 1. 2. 3. The Human Protein Atlas (HPA) is a research project that aims to visualize the cellular and subcellular location of every human protein by using monospecific polyclonal antibodies1. This high-throughput approach enables the discovery of new protein distribution profiles and can also provide deeper insights into already known patterns. Here we used HPA antibodies to identify proteins associated with the age pigment lipofuscin. Lipofuscin is mainly present in post mitotic cells. It consists of lipids, proteins, carbohydrates and a small amount of metals. Despite that lipofuscin has been well-known for a long time the exact composition of the protein portion is uncertain 2.

Fontana Sudan Auto- HPA025226 HPA041736 HPA045679 D-PAS Masson Black fluorescence HPA025226 HPA028498

Cerebral cortex 1. HPA025226, band in liver lysate around 40kDa (predicted mol weights are 36kDa and 18,4kDa.) 2. HPA041736, band in heart muscle lysate around 22kDa Heart (predicted mol weight is 17.7kDa.) muscle 3. HPA045679, band in heart muscle lysate around 26kDa (predicted mol weight is 27,2kDa.)

Liver Conclusions All methods; histochemical staining, confocal imaging and Seminal vesicle Western blot verified that the staining pattern observed by the antibodies was indeed lipofuscin. The large collection of antibodies produced within the HPA project has proven to be a valuable tool to identify compartments of Kidney cellular complexes like lipofuscin.

A number of the previously unknown proteins within lipofuscin have been identified; GGH (HPA025226), MIP18 (HPA041736) and Adrenal HDGL1 (HPA045679). Thereby a few pieces of the lipofuscin puzzle gland have been solved.

References Histochemical staining of lipofuscin using antibodies, D-PAS, Fontana Autofluorescent lipofuscin 1. A human protein atlas for normal and cancer tissues based on antibody proteomics. Masson and Sudan Black show similar results in all tissues. confirmed by immunofluorescence. Mol Cell Proteomics. 2005 4 (12):1920-32 2. Lipofuscin: Mechanisms of age-related accumulation and influence on cell functions. Free Radical Biology & Medicine, 33 (2002), pp. 611-619

The HPR project is funded by the Knut & Alice Wallenberg foundation. The atlas is part of www.proteinatlas.org the HUPO Human Antibody