This is Book Y, which begins Monday, March 9th, 1968, and ends somewhere towards the beginning of October. The first page is just a continuation of an experiment with one of the modified li‑‑ Bence Jones proteins, 26466, which have been derivatized with cysteamine. And continuing the purification of that derivative. And that continues on page 3 and ends on page 5, with the recovery of 196 milligrams of material. And the light chain, now, of 27489 [00:01:00] derivatives, beginning on page 7, similar sorts of work, of making and digesting of the light chains.
Rather heavily underscored, on page 11, Monday, March 18th, a time series of digesting — SS‑digest time series or cysteamine derivative time series. That’s the work that’s going on at that time, deri‑‑ digesting the material, the light chains, with TPCK trypsin — trypsin-derivatize‑‑ to be more specific.
A series in sodium bicarbonate, on page 11, [00:02:00] continuing with these digestions. April — light chain derivatives again, April 3rd, Wednesday, page 15. Some have been (laughs) lost, I guess, from the look of the comment on page 14. “Several are severely depleted. Found a leak in UM‑2 today. This has cause‑‑ Use only good ones.”
New digests, on the following day. And repeating digests continuing, as we go on through the book. [00:03:00] Different gradients, to make a better column fractionation. A new series of buffers for peptide analyzer. A couple of gels on page 26, of 8‑molar urea, formic acid gel, alpha purified light chains. And, “One [F(ab’)2?]?” question mark. (pause)
More digests. New digests, new derivatives, as we go through May now. Repeat [00:04:00] digests. So, many different digests of these various light chains. With a column reasonably well documented, on page 40 and 41, with the chart of a column and a gel of the product. And this following page, a similar thing, for [Phillips?], “[FI‑63 kappa crude?] clean-up” — kappa light chains — was the idea.
More work, on another Bence Jones protein that had been studied before, Dylan 2778 and 27932, repeats. [00:05:00] Etc. Trying to do dige‑‑ Bence Jones cleanup, on June 25th. More columns in the following pages. A lam‑‑ a lambda light chain here, being considered, on page 57 — lambdas Nevin and [Way?]. These are two light chains of Bence Jones proteins that are lambda rather than kappa.
Thursday, July 4th. “ID!” exclamation mark, Independence Day. Looking at the — what later were my favorite derivatives, those that have a [00:06:00] [SEtOH?] side chain on them — made with redu‑‑ dithiothreitol, a small amount, and a large excess of diethanoldisulfide, DDF, which is later on — was published as a very good method — which I think I’ve already referred to there.
We provisionally identified the extra SS peak in [1‑2 SA+?]. That’s a light chain variable, the light chain genetic variation. And [00:07:00] various diagrams.
Trying the ethylamine derivatives, on the 5th of July, chromatograms, which have faded rather extensively over time, on page 64 — continuing this work with these proteins. Trying to get the arginine reaction to work better, on page 69 — with a Technicon, [Auto-Technicon?] manifold. The Auto-Technicon was a big part of our tools, in those days, used for many different purposes. [00:08:00] Nothing remarkable happening here. August 7th, new derivatives. Preparative column, the following day.
Dansyl — dansylated amino acids being considered, on page 83, with quite an little image of separated dansylated amino acids, and the comment that “This system resolves alanine and valine.”
And continuing again. Quite pretty — quite pretty images to look at, of the [00:09:00] chromatograms of the various derivatives, page 85. Continuing. I ought to get some enjoyment of visualization of gels and chromatograms and so on. These are quite satisfying. “N‑terminals?” question mark, on page 89. More images, on page 91. N‑terminals again. Four or five images, on page 93, of derivatizing the amino acids. I must have been enjoying the… With, page 97, “The problem therefore is to retain chloroform/methanol resolution without a jagged front.” [00:10:00] Because I could see it wasn’t exactly straight, on the images to the left of that page. More chromatogram. Various solvents plus or minus water. Ninety percent chloroform, 10% methanol. Not very good. More pages of this type, 105, 106, 107, 108.
A full-scale test, on 109, with a large tank. A frame in which there was a — chromatograms — which are quite big-scale. These were large piece‑‑ large re‑‑ pieces of filter paper. And the image on the left, on page 108, is [00:11:00] quite satisfactory, with a comment, “The shapes suggest no evaporation” — with a mixture of 90 parts chloroform, 10 parts methanol, and three milliliters of proprionic acid. More of this type of result, on the following pages, trying to learn how to do chromatograms with amino acids, derivatized. Pages and pages of this sort of thing.
By page 121, I’m now trying controlled evaporation, to look how it changed the shape of the chromatograms, which is — quite obviously did change them quite substantially — with various little devices to try to improve the method.[00:12:00] Attempt with a large tank, straight, and tested, on page 123 — but with a comment that “Something’s very wrong, (laughs) relative to Y‑113. Almost no movement [of?]‑‑” on the left-hand page. Although the chromatograms don’t look bad. More of the same type of thing, as the book progresses, nothing terribly exciting.
On page 129, “Even more” use‑‑ “more careful saturation is desirable. Cover the top with felt?” question mar‑‑ “And soap.” Various conclusion. “No detectable alanine.” [00:13:00] But, “#12 –” looks like “no leucine is OK” — etc., etc. — as this book is drawing to a close.
Propanol system, October. N-propyl alcohol, page 135. More chromatograms, and other sheets, Sunday, October the 6. I probably enjoyed doing these. Quite a lot of them, [they’re?] visually, as I said, quite satisfying. Methanol, acetic acid, and chloroform, on page one-one-thr‑‑ 143, as we get to the end. N‑terminals, [00:14:00] on Tuesday, October 9. (pause) As the book draws to a close. October 9, N‑terminals again. And ends on page 152, with another pair of chromatogram, not very revealing, at this point in my life. End of Book Y.