Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA035113

Local Sample ID:	GTM14L1
Subject ID:	SU000647
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Species Group:	Human

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Sample Preparation:

Sampleprep ID:	SP000654
Sampleprep Summary:	Isolated trabecular meshwork tissue was subjected to an alternating cycle of immersion in liquid nitrogen and 37˚C, followed by extraction of lipids using a modified Bligh and Dryer method.13 The organic phase with extracted lipids was dried in a Speed-Vac (Model 7810014; Labconco, Kansas City, MO). Samples were flushed with argon gas to prevent oxidation. Corresponding aqueous phase extracted proteins were subjected to determination of concentration using Bradford’s method. A subset of protein samples also were subjected to densitometric quantification using bovine serum albumin as a standard (amino acid quantified) after electrophoretic separation on a PHAST (GE Healthcare Bio-Sciences AB, Uppsala, Sweden) gel system. All extractions and subsequent handling were made using glass vials, and polyvinyl plastic was avoided completely. We also added a PC control standard during tissue homogenization, determined its recovery in an aliquot, and used for the calculation of total recovery of this standard for each extraction to ensure >99% recovery of added standard during extraction.

Sampleprep ID:

SP000654

Sampleprep Summary:

Isolated trabecular meshwork tissue was subjected to an alternating cycle of immersion in liquid nitrogen and 37˚C, followed by extraction of lipids using a modified Bligh and Dryer method.13 The organic phase with extracted lipids was dried in a Speed-Vac (Model 7810014; Labconco, Kansas City, MO). Samples were flushed with argon gas to prevent oxidation. Corresponding aqueous phase extracted proteins were subjected to determination of concentration using Bradford’s method. A subset of protein samples also were subjected to densitometric quantification using bovine serum albumin as a standard (amino acid quantified) after electrophoretic separation on a PHAST (GE Healthcare Bio-Sciences AB, Uppsala, Sweden) gel system. All extractions and subsequent handling were made using glass vials, and polyvinyl plastic was avoided completely. We also added a PC control standard during tissue homogenization, determined its recovery in an aliquot, and used for the calculation of total recovery of this standard for each extraction to ensure >99% recovery of added standard during extraction.