BPA stimulates proliferation in JKT-1 cells through a G-protein–coupled receptor (GPCR). GPCRs have been proposed to be involved in triggering membrane action of steroids ( Thomas et al. 2006 ), including estrogens ( Filardo et al. 2002 ; Kelly and Wagner 1999 ). PKA activation is usually a Gα s protein-dependent mechanism, so we tested the effect of NF449, a Gα s inhibitor ( Figure 6 ). NF449 blocked the promoting effect of BPA, illustrating the Gα s dependence of the PKA activation and the G-protein–coupled nature of the receptor involved in BPA stimulation. We have previously shown that E 2 -BSA, which triggers an effect quite similar to that of BPA, also induces a Gα i -dependent activation of the mitogen-activated protein kinase (MAPK)/ERK1/2 pathway ( Bouskine et al. 2008 ). For this reason, we also studied the effect of PTX, an inhibitor of Gα i /Gα q protein, during BPA-induced JKT-1 proliferation. This toxin prevented the BPA-induced increase of cell proliferation ( Figure 7 ).
Steroid isolation , depending on context, is the isolation of chemical matter required for chemical structure elucidation, derivitzation or degradation chemistry, biological testing, and other research needs (generally milligrams to grams, but often more  or the isolation of "analytical quantities" of the substance of interest (where the focus is on identifying and quantifying the substance (for example, in biological tissue or fluid). The amount isolated depends on the analytical method, but is generally less than one microgram.  [ page needed ] The methods of isolation to achieve the two scales of product are distinct, but include extraction , precipitation, adsorption , chromatography , and crystallization . In both cases, the isolated substance is purified to chemical homogeneity; combined separation and analytical methods, such as LC-MS , are chosen to be "orthogonal"—achieving their separations based on distinct modes of interaction between substance and isolating matrix—to detect a single species in the pure sample. Structure determination refers to the methods to determine the chemical structure of an isolated pure steroid, using an evolving array of chemical and physical methods which have included NMR and small-molecule crystallography .  :10–19 Methods of analysis overlap both of the above areas, emphasizing analytical methods to determining if a steroid is present in a mixture and determining its quantity.