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It may be worth way more than 2 cents. Yesterday when I was adjusting pH's of the buffers I realized that starting at about pH 8.2 and up this mix is not really a buffer. pH was changing very fast with little of solution being added. There are number of buffers (including Good buffers) that will cover pH's between 3-11. One place to look is a Hampton "pH Stock Option". The intriguing thing was to read in several papers where authors claimed that crystals could be obtained only in Imidazole-Malate buffer and including pH 8.5. Thank you. Vaheh Oganesyan, PhD Scientist II MedImmune, Inc. One MedImmune Way Gaithersburg, MD 20878 USA Phone: (301)398-5851 Facsimile: (301)398-9851 www.medimmune.com -----Original Message----- From: Nadir T. Mrabet [mailto:[EMAIL PROTECTED] Sent: Wednesday, January 10, 2007 6:19 AM To: Oganesyan, Vaheh Cc: [email protected] Subject: Re: [ccp4bb]: Imidazole-Malate Buffer My 2-cents worth. All pKa are given at 20 °C. pKa (malate ; pK2) = 5.13 pKa (imidazole) = 6.95 Hence, usefull buffer range goes from 4.13 to 7.95 with better buffer capacity from 4.4-7.7. I wouldn't dare using the mixture as a buffer beyond pH 7.7 and certainly not at pH 8.5. If you need to cover the pH 5.5-8.5 range, I would suggest using an equimolar mixture of MES (pKa = 6.15), MOPS (pKa = 7.2) and HEPPS (pKa = 8.0), all in acidic forms with titration with KOH (the amount of which can be exactly calculated using the Henderson-Hasselbalck equation to yield the desired pH value, in a highly reproductible fashion). Furthermore, using switterionic buffers, you no longer have to worry about activity coefficients as is the case with malate. Good luck, Nadir Mrabet Pr. Nadir T. Mrabet Cellular & Molecular Biochemistry INSERM U-724 UHP - Nancy 1, School of Medicine Avenue de la Foret de Haye, BP 184 54505 Vandoeuvre-les-Nancy Cedex France Phone: +33 (0)3.83.68.32.73 Fax: +33 (0)3.83.68.32.79 E-mail: [EMAIL PROTECTED] Cell.: +33 (0)6.11.35.69.09 LEGAL NOTICE Unless expressly stated otherwise, this message is confidential and may be privileged. It is intended for the addressee(s) only. Access to this E-mail by anyone else is unauthorized. If you are not an addressee, any disclosure or copying of the contents of this E-mail, or any action taken (or not taken) in reliance on it, is unauthorized and may be unlawful. If you are not an addressee, please inform the sender immediately. Oganesyan, Vaheh wrote: > *** For details on how to be removed from this list visit the *** > *** CCP4 home page http://www.ccp4.ac.uk *** > > > Thanks to Marc Graille, Robyn Stanfield, Artem Evdokimov and Pietro > Roversi for sharing their experience on how to make Imidazole-Malate > buffer. > Essentially it is same as making phosphate buffer. In the range of pH's > from 5.5 to 8.5 mix the un-adjusted d-Malic acid and Imidazole solutions > in > specific proportions to get desired pH. > > Regards, > > Vaheh Oganesyan, PhD > Scientist II > MedImmune, Inc. > One MedImmune Way > Gaithersburg, MD 20878 > USA > Phone: (301)398-5851 > Facsimile: (301)398-9851 > www.medimmune.com > > -----Original Message----- > From: Pietro Roversi [mailto:[EMAIL PROTECTED] > Sent: Tuesday, January 09, 2007 4:33 AM > To: Oganesyan, Vaheh > Cc: [email protected] > Subject: Re: [ccp4bb]: Imidazole-Malate Buffer > > Dear Vaheh, > I usually prepare the 2M solutions and then: > > 1. If a final basic pH is needed, I add malic acid to the imidazole > solution until the desired pH is achieved > 2. If a final acidic pH is needed, I add the imidazole to the malic acid > solution until the desired pH is achieved > > With best wishes > > Pietro > -- > Pietro Roversi > Sir William Dunn School of Pathology, Oxford University > South Parks Road, Oxford OX1 3ER, England UK > Tel. 0044-1865-275385 > > >
