Journal:

Article Title: HS1 protein is differentially expressed in chronic lymphocytic leukemia patient subsets with good or poor prognoses

doi: 10.1172/JCI24276

Figure Lengend Snippet: 2-DE proteomic analysis of purified CLL cells. (A) The square identifies 2 close protein spots with the same Mr of 79 kDa and different pIs, 4.83 and 4.86, which were identified as HS1 proteins by MALDI-TOF MS analysis (see Results). IEF, isoelectric focusing. (B) Both right and left protein spots were expressed in 2D gels obtained from 2 representative CLL patients with good prognoses (subset 1) while only the more acidic one was present in 2D gels obtained from 2 representative CLL patients with poor prognoses (subset 2). Arrows indicate the left and right protein spots identified as HS1. (C) Proteins obtained from 5 representative CLL patients were resolved on 2-DE that was either silver stained (left panel) or transferred onto nitrocellulose and incubated with an anti-HS1 antibody (right panel). One representative case is shown. The antibody hybridizes in an area corresponding to the protein spots of interest. WB, Western blot.

Article Snippet: The pellets were lyophilized and solubilized with 2-DE buffer (9 M Urea; 10 mM Tris; 4% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate [CHAPS]; 65 mM DTT; 2% IPG buffer ampholine, pH 4–7 [Amersham Biosciences]; and protease inhibitor cocktail).

Techniques: Purification, Staining, Incubation, Western Blot

Journal:

Article Title: HS1 protein is differentially expressed in chronic lymphocytic leukemia patient subsets with good or poor prognoses

doi: 10.1172/JCI24276

Figure Lengend Snippet: The more acidic protein spot is a phosphorylated form of HS1 protein. Cells from 4 CLL patients belonging to subsets 1 and 2 were treated with λPPase; proteins were resolved on 2-DE, transferred onto nitrocellulose, and incubated with a specific anti-HS1 antibody at different time points. One representative case is shown for each subset. The densitometric analysis indicates the progressive shift of the HS1 protein from a more to a less acidic form after 7 and 15 hours of incubation.

Article Snippet: The pellets were lyophilized and solubilized with 2-DE buffer (9 M Urea; 10 mM Tris; 4% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate [CHAPS]; 65 mM DTT; 2% IPG buffer ampholine, pH 4–7 [Amersham Biosciences]; and protease inhibitor cocktail).

Techniques: Incubation

Journal:

Article Title: HS1 protein is differentially expressed in chronic lymphocytic leukemia patient subsets with good or poor prognoses

doi: 10.1172/JCI24276

Figure Lengend Snippet: HS1 pattern of expression in normal B lymphocytes. (A) Purified tonsillar B cells were cultured in the presence or absence of anti-IgM antibody for 2 minutes. Proteins from untreated and treated cells were resolved on 2-DE and revealed by silver staining. The arrows indicate the 2 HS1 protein spots, and 3D-densitometric analysis (right panels) shows a shift from a less acidic to a more acidic form of HS1 protein following BCR stimulation. nc, negative control. (B) Proteins obtained from FACS-purified naive and memory tonsillar B cells were resolved on 2-DE and revealed by silver staining. The top panel of each B cell population identifies the different HS1 forms. The 3D-densitometric analysis shows that both protein spots are expressed in similar amounts in naive B cells, while in memory B cells, the more acidic form is predominantly represented.

Article Snippet: The pellets were lyophilized and solubilized with 2-DE buffer (9 M Urea; 10 mM Tris; 4% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate [CHAPS]; 65 mM DTT; 2% IPG buffer ampholine, pH 4–7 [Amersham Biosciences]; and protease inhibitor cocktail).

Techniques: Expressing, Purification, Cell Culture, Silver Staining, Negative Control