Ammonium sulfate saturated

Treatment of the culture filtrate with increasing ammonium sulfate amounts showed that the bulk of polygalacturonase activity precipitates between 45 and 85 % ammonium sulfate saturation (Table 1). 

Bring filtrate to 70% ammonium sulfate saturation (472 g ammonium sulfate/liter filtrate), adjust pH to 8.0 using sodium hydroxide, and stir for 1 hr. 

Incubate all the tubes on ice for 20 min. NOTE It is very important that the protein solution be brought up to the desired ammonium sulfate saturation slowly. If this step is not done correctly, it will be difficult to reproduce these results when the same procedure is carried out on the bulk preparation. Also, avoid frothing in the solution (a sign that some of the proteins are being denatured) by gently mixing the solution as the ammonium sulfate is added. 

Calculate the concentration of protein (in milligrams per milliliter) present in FR-I, FR-II, and each of the ammonium sulfate saturation trials. 

Prepare a plot of percent GOT recovered versus percent ammonium sulfate saturation using the data from tubes 2 to 7 with the use of the following equation ... 

Determine the highest percent ammonium sulfate saturation that gave less than 20% GOT recovered. This will be a good percent ammonium sulfate saturation to use for the first cut on Day 3 (when the bulk of the GOT is still soluble). Next, determine the lowest percent ammonium sulfate saturation that gave more than 80% GOT recovered. This will be a good percent ammonium sulfate saturation to use for the second cut on Day 3 (when the bulk of the GOT will precipitate out of solution). This information will he critical for the experiment performed on Day 3, so these calculations must he completed prior to the beginning of the next experiment. 

Remove the supernatant from the precipitated IgG pellet in each tube and discard. The serum consists of 80 to 90% albumin, which is soluble at 50% ammonium sulfate saturation. The IgG is insoluble at 50% ammonium sulfate saturation and will precipitate. 

Jacob et (d. (Jl) used 4-step ammonium sulfate fractionation of hog gastric mucosal extracts, followed by ultracentrifugation, to obtain high-potency intrinsic factor-active materials (Table 5). The intrinsic factor-containing fraction precipitated between 39 and 47% ammonium sulfate saturation was active at a 1-mg level. About 90% sedimented with a constant of 1.78-1.82 S. Other heavier components formed only about... 

Although as many as eight distinct proteins have been demonstrated immunologically in the soluble fraction obtained at 65% ammonium sulfate saturation (2.6 M), the total amount of these antigens represents a very small part (about 3%) of the total albumin fraction (06). 

Ammonium sulfate precipitation. Mixing of the ammonium sulfate with the antibody solution is not instantaneous and takes time, which depends on the properties of the components as well as on the processing conditions, i.e., mixing intensity, the volume, density and viscosity of the medium as well as on the power input for mixing. The volume of saturated ammonium sulfate solution in mL that has to be added to a sample to salt out proteins at any concentration is given by the equation V (mL) = 100 mL X (S2-S1)/(1-S2) where SI and S2 are expressed as fractions of the saturated solution, 100 is the volume of the sample and V(mL) is the volume of saturated ammonium sulfate that has to be added to the sample to obtain the desired concentration. For example A 100 mL sample is precipitated sequentially first to an initial saturation of 25% and then to a final 50% ammonium sulfate saturation. V(mL) = 100x(0.25-0)/(l-0.25). V(mL) = 33.3 mL. The volume of 33.3 mL of saturated ammonium sulfate is added... 

The isolation of protein from seaweeds is a difficult task due to the link between polysaccharides and protein within the seaweed matrix. It is described that the extraction of proteins from the tissues of laminarialean algae (Nagai et ah, 2008) or Saccharina japonica (Kim et ah, 2011) is difficult due to high levels of nonprotein interfering compounds, mainly viscous polysaccharides. As a consequence, isoelectric point (Ma et ah, 1996) or ammonium sulfate saturation (Hemandez-Mireles and Rito-Palomares, 2006) or trichloro acetic acid (Barbarino and Louren o, 2005) approaches, which are cormnonly used for protein precipitation, are not completely useful for seaweeds. Thus, to solve this task, different approaches are proposed such as proteolytic treatment of the whole seaweeds followed by filtration and dialysis (Suetsima and Nakano, 2000) or treatment... 

The processing activity can be selectively precipitated by ammonium sulfate and is sensitive to boiUng. The processing enzyme is precipitated between 40% and 70% ammonium sulfate saturation of the crude extract. The enzyme is inactivated if the extract is boiled for one minute, whereas mock reactions without added extract show no specific cleavage or protein degradation. 

The nucleosides obtained by enzymatic digestion of t-RNA were separated by bidimensional chromatography using Merck cellulose layered on 20 x 50 cm aluminum sheets. The elution in the first direction was performed with n-butanol-isobutyric acid-ammonium hydroxide, conc.-water (75 37.5 2.5 25, v/v) mixture for 24 hr at 20-21°C, in an unsaturated chamber. The chromatographic plate was dried with a fan for 3 hr at 20°C, after the first elution. Elution in the second direction was performed for 4 hr with the mixture ammonium sulfate (saturated solution)-0.1 M sodium acetate (pH 6)-isopropanol (79 19 2, v/v). The separated compounds were visualized in UV light at 254 mn. 

From Na-deoxycholate solubilized brain microsomes the fraction that precipitates between 45 and 60 per cent ammonium sulfate saturation was obtained. This fraction contains ATPases that can be activated with Mg2+ or Ca2+ or Mn2+ in the presence of EDTA and ouabaine (Table 4). In the presence of EGTA (ethanedioxy bis (ethyl amine tetra-acetic acid)) but not ouabaine, and Mg2+ the addition... 

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