Oliver Smithies:
— book, Roman XV, 15, November 1957 through January 1958, and it’s basically a compilation of a lot of data. I think we won’t find anything terribly exciting in it, just data. First significant entry is on November 19th, where there are a whole bunch of tests with different photographic procedures, KB14 film, D8, for three minutes, on F2 paper, etc., that sort of thing. And different films being tested — I mean, different exposures being tested. [00:01:00] Tested multiple, one-dimensional D8 three-minute microfile with red, no, light green, dark green, and blue filters. Red is approximately equivalent to [non?], but the contrast is still too great. The contrast is still great, and trying to find a filter that would allow a record without being fully saturated, either on the white end or the black end, with a note, the KB14 one second with F11 is approximately correct, and KB17 one second at F16, and followed the next page by some two-dimensional gel data, [mini?] gel.
[00:02:00] On November 21st, two-dimensional and one-dimensional gel, showing that, in fact, under some circumstances, double beta can be seen in a one-dimensional gel. Not very beautiful result, but they are there. Here’s a gel here, a large gel with multiple slots, Saturday, Sunday, November 24. The slots were full. [00:03:00] (laughs) (inaudible) seven volts per centimeter, and very much too hot gel, though the volts per centimeter went down to about four or five. Nonetheless, I note that I could see three very clear beta BC — in other words, the double transferrin. Later on, that sort of comparison is needed in order to test differences in the beta globins. When they have to be alongside each other, two dimensions is not very good for that. For example, on Sunday, Monday, November 25th, I have a comment that the beta BC is in two of the samples, but [00:04:00] possibly not equivalent. This is one-dimensional electrophoresis. Just looking at the sample there. Two are alike and one is a little different, but it’s rather faint. That may just be a concentrating effect, but I didn’t — I just was alert to the fact that there might be more than two variants of the beta globin. More films with and without hemoglobin, multiple slots, multiple samples. So these are large gels with multiple slots in the gels, that are still being run horizontally, I think, at this point, though we’ll find out in a little while. [00:05:00] Two-dimensional gels, Tuesday, November 26th. Nothing very remarkable. Continuing in this vein. And comparing two-dimensional and one-dimensional, November 28th. Repeat of two-dimensional to check BC versus CD, beta BC versus beta CD. With a comment that disregard all doubles [00:06:00] but the three. Three are one-dimensional from BC. I’m not sure what that means. Had a water cool gel here, on Friday, September — Friday, Saturday December 7th. Six volt per centimeter, wide slots, water cooled, five centimeter depth, whatever that means. (laughs) Convection, circulation. I must have been immersed in water, five centimeter deep, and allowed it to cool just by convection. The result is really quite nice, very pleasing results, the curvature almost zero, but [00:07:00] really, looking at the result and the comment, it is really quite a nice gel. Still horizontal, I think.Monday, Tuesday, December 10th, similar sorts of experiment, of the two-dimensional confirmations on Tuesday, December 10th. (laughs) [One here?] that I call it on Tuesday, Wednesday, December 11th, on the date, [5:00 p.m.?], six volts per centimeter, with a self-slicing tray. (laughs) I don’t know what that was, but the results are quite nice. A bit heavy. [00:08:00] Nine volts per centimeter, cool gel on Thursday, Friday, December 13th. Not very good. A large number of samples here that are presumably cattle samples, and just labeled with numbers 6,000, 6,147, etc., 6,842, 6,970, different samples, but no image there. Friday, December 13th, the setup of 4:00, 5:00 p.m., [00:09:00] 11 volts per centimeter, cooled with methanol in this case, and a comment that the beta and the hemoglobin haptoglobin type one have resolved, which I never before succeeded in doing in a one-dimensional gel. Try increasing the volts per centimeter to 50 with efficient cooling, and two millimeter self-splitting gel. I think the self-splitting gels were probably gels cast once, and then recast on top of them. I’m not quite sure what they were. Methanol air-cooled on Saturday, December 14th. [00:10:00] Some contamination noted between the gels, between the sample. It’s fairly obvious from the image, rough handling during the setting up is the cause.
Check of aging sample. So water cooled — water air cooled, 12 volts per centimeter, December 16th, higher volts per centimeter, difficult to keep the temperature down with the water 20.5 degrees centigrade, fairly stable [00:11:00] at high air, [but?] water air cooled, trying to get high voltages. Prepared a sample. The gels are quite nice. You can see the resolution between that albumin and beta globin is good.
Still worried about what I call beta splitting. Set up two-dimensional… And water air cooled bovine samples, really quite pretty, on Tuesday December 17, 12 volts [00:12:00] per centimeter, water air cooled, the temperature increased to 23 degrees, so I added some methanol, but the beta globin bands are beautifully resolved in these gels. Water air [methyl?] cooled, 20 volts per centimeter. Not very good. Results poor. These are still definitely horizontal gels, but they are quite good results. The slots are open, I think. [00:13:00] Twelve volts per centimeter again, here looking at the resolution — this is December the 19th — looking at the resolution of the bands between beta globin and albumin, but also looking at the transferrins. Results good, but better at 20 degrees. These were 25 degrees. So try 15 volts per centimeter at 20 degrees, so working to get better results by cooling and having high voltages is the motivation [00:14:00] of these experiments. Still some two-dimensionals in there, in December 20th, 22nd.
So these are Friday, December 20th. It’s fairly clear that I haven’t really quite understood what the gels were showing, because in the notation of beta globins, because on December 20th the sample shows really that the double ones are transferrins, that is faster, not slower, [00:15:00] than the standard one, and this is with what I call beta BC. I must have had beta BC and beta BD and not realized it in talking about the samples earlier. This is of transferrin or beta globin still, as far as I knew at this point, faster than the normal one versus one that’s slower. Not possible, really, to tell in the two-dimensional gels. They have to be compared in one-dimensional gels to be sure. For example, Saturday, December 21st, there’s a two-dimensional gel with [Rex Thompson?], and it would appear that the [00:16:00] double band is due to the common one, and one that is slower, Rex Thompson’s. And yet, looking at the December 20th gel, the one-dimensional gel, it’s clear that Rex Thompson — one, two, three… No, it’s not clear. No, I’ve misinterpreted these results. I’ll go back to this and start that part again.
Looking at Friday, December 20th, some one-dimensional gels, looking at various samples, in order to [00:17:00] understand, again, what variations there are in the beta globin, and I have these sample 5879 III 2297269, and 5865 VI. These are samples where the variant of beta globin is faster than the common one, not slower, whereas it would appear that perhaps Peter [McGuigan?] has a double band with a slower one, as well. Not completely clear. But looking then at the three-dimensional, two-dimensional gel, [00:18:00] on Saturday, December 20th, one can see that Peter McGuigan is a single band, not a double one, and that Rex Thompson has a double band. And it would appear there to be slower, but, in other words Rex Thompson being slower, but on the… It’s still confusing. Rex Thompson looks to have a slower band. And Rex Thompson, on the December 20th, one-dimensional gel, has a single band, so it’s [00:19:00] confusing. There’s something not quite right here. Presume it’ll be sorted out.
Tried some 6 volt per centimeter [overnight?] gels with hemoglobin, and a comment, not suitable. But quite sufficient to resolve beta BC. That’s a faster sample, faster beta. It’s just my not remembering that has caused the confusion, not really the data. The data eventually are quite clear that you had various forms of [00:20:00] transferrin. Still two-dimensional, [mini?] two-dimensional gel. A whole bunch of samples obtained January 3rd, 1958, referring back to two previous books, Roman 23, Roman 24, which [00:21:00] I have some answer to why that’s written… It’s not my writing. I think that probably is book 13 and book 14, not book 23 and 24. Might be. January 3rd, 1958. No, that’s still the same book. It probably means book 13 and book 14. Photographic tests, still, with microfile, always, constantly fiddling with these variables. More two-dimensional gel. Some one-dimensional gel. Again, two — also, Monday, Tuesday, January 7th, just looking at haptoglobin [00:22:00] types in these, basically. Whole bunch more samples, still with this 25 and 26 and 27 notation, all in Otto’s writing, I think.
Thursday, Friday, January 10th, trying to improve the [00:23:00] post-albumins, as I called them, 95% full slots to improve slow alpha 2, and ran them horizontally. 2A, 2B, and all three haptoglobin types are in there. The resolution of the post-albumins is not particularly good, but doesn’t say anything about it in the notebook. So I could comment on the difficulty of making tape, slot form, tried some different forms of slot form. [00:24:00] Worried about electrodecantation, which is common with an open slit, and a sample with nothing in support. Electrodecantation was always a worry in running horizontal gels, had been the problem all along. And a tapered slot being tried, 1 and 10 taper on Monday, January the 13th, the top and the bottom of the gel, and various [related?] comments. All showed electrodecantation, very bad, but reversed with the gamma. In other words, the gamma is — [00:25:00] goes up when the serum goes down. Still haven’t made the transition to a vertical gel yet. Tuesday, Wednesday, January 15, just a simple, multiple-slot horizontal gel with open slots.
[00:26:00] Just running samples for typing without any particular comments about them in the following pages. Typical being Thursday, Friday, January 17th, six volts per centimeter, self-slicing gel, and the results from it, typing the different haptoglobin, a comment on the transferrin of the beta globin of one sample. Quite good results, January 18th, Friday, Saturday, [00:27:00] not very good, but quite good. And that takes us with Monday, Tuesday, January 21st, six volts per centimeter, etc. Nothing particularly remarkable, but the end of this book.