AFM: one limitation affecting peak number of SP-D dodecamers

AFM: one limitation affecting peak number of SP-D dodecamers, could be what I found on a website. “Due to the nature of AFM probes, they cannot normally measure steep walls or overhangs“. This quote is from a website that is mostly in chinese so I didnt bother to read or translate. BUT it is clearly what I have seen as I search for peak number in SP-D molecules (whether trimers, hexamers dodecamers, or higher order multimers).

Case in point is how many times the “tiny peak” which I have noted dozens and dozens of times as a small in height, narrow peak right near the base of the N term peak. It seems to be missed a lot of the time, there are other times when it is very obvious. A couple reasons for its intermittent appearance in AFM could actually be because of the very steep wall on the downslope of the N term peak before the glycosylation peak.

In terms of “overhang” this might also apply to the blending and disappearance of the neck area which is sometimes covered in the CRD peaks which flop around visibly lying over the neck domain.

Quilts forgotten:

This will be a different kind of a post. It reflects my own “terror” of being old and “being forgotten” maybe even more, the total loss of anything I did in my life as meaningful. I have tried to be kind, thrifty towards myself, generous to my family, and others, but nothing I have done would seem to have had much impact on the planet, on life, on the present, or on the hereafter, whatever the hereafter is.

I like to quilt. I quilt with scraps, tiny scraps (down to 1.5in square). Its like putting together something organized out of something lost in entropy, destined for land fill in some foreign country, saving grief from the world wide land fill.

I like putting the scraps of “nothing” together into something orderly. I don’t quilt all the time, incessantly, but about one or two quilts per year, using my old (now ailing) green metal singer sewing machine. Many quilts I made had clothes from my three children, boy scout color light brown blocks from their shirts, and pink glossy fabric from my daughter’s play dress up dresses. Fabrics from PolyFlinders left over boxes of cutter’s scraps… 7 dollars for several pounds of stacks of cute ginghams, flower prints, delicate little girl fabrics and sometimes yard goods, ribbons, thread etc. (They were on 8th street i believe, in cincinnati in the 1980s). I enjoy quilting.

I have a colleague from UC who left the university long before i retired, but he offered me a bag of fabric scraps, since he knows I quilt with scraps. These scraps were presumably from a son in law whose 93 yo mother had fallen and was going into health care.  It was a heavy bag of scraps, I could see from the outside that some piece work was there.

I was not prepared for what was inside. Nine (9) finished quilt tops, varying sizes, all waiting to be quilted into bedspreads, or coverlets.  I was full of grief: for that woman who’s last dozen years were spent hand quilting these precious gifts for her children, grandchildren and friends and they were never finished, never given as treasured gifts, and then “pitched” to an unknown friend of a friend, as if the thousands of hours of hand piecing these quilt tops was “nothing” “nada” “rien” “zilch” “zero” “trash”.  It grieved me so much.  They will go back to the giver. Hopefully with instructions on where to find someone who will add batting and a back, and finish what this woman (who is still alive), can give her children and friends.

There were three quilts i suspect were for three little boys, in particular, maybe one for her grandson, they looked pretty much alike. I hope when i return them, they are finished by a professional and given to individuals, just where they were intended, as finished works, they can ask her who was to get which.

I was totally grieved.  For her yes: for myself yes, as I know beyond hope, that because I have not completed my own grandmothers, necktie quilt, whose silk is shreding from a 100 years of age, that the quilts i have made will be similarly “pitched” as will my grandmothers quilt.  I dont find those thought entertiaining at all, i find them totally depressing.  And in fact, there is nothing new in my thoughts….. dust to dust and there is nothing then to remember, to treasure, to feel proud of, to hand out as a legacy. That is just how this journey works.   See pix below…. all 9 of that grandmother’s hand work, tossed, but maybe saved and given as gifts.

So this also begs the question, why were these quilt tops NOT finished. I suspect the reasons are just like my quilting. Not perfectly straight, there will be lumps in the quilting with puckers in the fabric, no straight corners, some colors showing through to the front. The backing and quilting the whole quilt is really a hard part.  I know. I have one pieced currently waiting to be finished.

Kudos to this woman, whose fingers didn’t stop when she got old.

Bobbin, sometimes called boobin…LOL

Bobbin, sometimes called boobin…LOL

While checking to find the part number for the motor belt on my old Singer sewing machine, i saw this typo for the bobbin winding belt.

This is one of the funnier typos online that i have found recently.  And no, Boobin is not the way bobbin is spelled in german.  I checked.

Surfactant protein D, peak plots from AFM images

Last couple months I have been plotting individual trimers of SP-D to determine if number of peaks, peak heights, a valleys using the same 6 methods used on dodecamers produces the same results. The same 6 functions were use on trimers as on dodecamers (See list of image and signal processing functions used — at the end of this post).

There is clearly a smaller N term peak (N) in the trimers than the dodecamers where four N term peaks are “piled” or “linked” together. Other peaks do not change much, which is a good thing.

The plot immediately below shows just the peaks plotted for just one trimer (kind of a perfect one), and immediately below that plot shows mean peak height, width, and variation for 6 dodecamers (24 trimers, 368 trimer plots),  and the third plot down is 14 dodecamers (56 trimers, 897 trimer plots) and the line graph below that is the plot from Arroyo et al, 2018 in which she identifies the N peak (peachie orange), the Gly peak (light green), and the CRD peak (orange) ONLY. My purpose was to show that there are at least 8 prominent, consistenly found peaks per trimer, not just three.

So many different images analyzed from different publications with Arroyo as author, are shown, ploted as dodecamers (6 or 14 individual dodecamers) or single trimer, there seem to be 8 peaks per trimer. Widest peaks are indeed the N, Gly and CRD, next widest and most consistend are peaks 4 and 6, the yellow peak (coiled coil neck domain) is infrequent, and during preparation for AFM is typically covered by the CRD domains flopping around, and also infrequent are the small peak called “tiny” because it is both thin and low (but consistently on the down slope of the N term peak) and peak 4 which is broad and rolling like the GLy peak, and a low and thin peak (pink), and a broad low peak before the coiled coil neck.

So after more than a thousand plots, I think I am pretty sure these peaks are part of the molecule and can perhaps contribute to creating a complete protein model of SP-D (which do date has not been done).

https://imagej.nih.gov/ij/
https://terpconnect.umd.edu/~toh/spectrum/PeakFindingandMeasurement.htm#ipeak
https://terpconnect.umd.edu/~toh/spectrum/PeakFindingandMeasurement.htm#findpeaksx
https://terpconnect.umd.edu/~toh/spectrum/PeakFindingandMeasurement.htm#Spreadsheet (s)
https://docs.scipy.org/doc/scipy/reference/generated/scipy.signal.find_peaks.html
https://stackoverflow.com/questions/22583391/peak-signal-detection-in-realtime-timeseries-data/22640362#22640362 (version: 2020-11-08)

Verge of a Dream: Two letters

There were two letters
You had never shown
To me. Two letters
On the back of a
Charm I guess
You’d say. I don’t know
The names of ornaments.
Of gold pieces worn on
A gold string of links.
A rosette fairly finely
Made with two letters
On back of the
Rose to
Lay
flat below the
Neck and above
All, the heart.
The letters
Began a name.
It is too clear
To ask how
Despondency
Comes from
The heart with
Sense unengaged.
The name,
from
The two letters
Engraved upon
A heart that
Was never shown to me.

RLB 02-08-2024

Verge of a dream: from that day forward

At work everyday,
almost, the
Tie you bought
I wear
Though there is
No reason to
Believe but it is hope
Which Has no equivalent.
That you will like
me more.
A clip for the
Tie, I’d ask for
Just ask, nothing
Is less useful
Than to beg but
It must be
Unquestioned of
Weight and
Solid gold.
So pure no one
Would try and
Scratch their
Way between us.
Or a pearl shirt
I wanted,
For you to
Give
made from
finest cloth
And though from
The loom it
Was unmarked.
I wash anyway
The bleach
Undoes the past
And there
my care
is sure that
You may like me
each fresh warm
tropical morning
Worn for you
From that day forward

RLB 02 06 2024

Verge of a Dream: How I met you

Near the block where
I grew up
I don’t know where I’d
Meet you
Across the gym
In your prom dress
With someone
Else
At work knowing
More than I could
Hope to.
I don’t know where
I’d meet you.
Without asking since
Asking brings rebuffs
Though they’re little
As if in a cart
Marked down.
Every one is sweetly
Given, no comment
of reservation,
Then, maybe to
Remember, you play,
The notes
Reminders each of
A moment between
Uncertainties that
Was happy. Now
You are not unhappy
But not like me
Blessed and blissful
In the moments
Remembering how I
Met you.

RLB 1/29/2024

Peak finding functions

Peak finding functions are certainly not the end all and be all of assessing peaks in AFM images of surfactant protein D. I am totally disappointed in how they work.  It is so clear that depending upon measures that have preceeded a peak and how big a peak is has LITTLE at all to do with what nature does with proteins (specifically, irregularly placed bumps and twists of varying sizes that can be seen by eye with electron microscopy and AFM and other microscopic media). Not a single peak finding function that I have found has any real comparison to what a trained eye can find (at least at this time). Since the user determines the robotic approach, peaks can vary from 4 to 40 when the eye can see about 15 in a dodecamer regularly.

When I see the divisions of peaks, the numbered peaks, the missed peaks that signal processing produces i think to myself. Why that, or why not this.

Sadly this makes everything I have tried to figure out about unbiased counting, height, width and valley depths, using image processing and peak finding, in surfactant protein D trimers (and dodecamers) is pretty much NOT verifying anything except the fact that they dont work.

Being adaptable mentally just is still a human trait……. how much longer, I dont know. Training seems still to lag behind human abilities.

Things I have noticed:

1– in the trimers, it appears that multiple peaks clustered (maybe 2 or 3) seem to happen when there are (as in the case of the glycosylation site (published data) the three trimers at the glycosylation site have shallow staggered peaks, as if there is a rotary positioning. Seems logical that the lumpy effect would be found when rotating a repeating event (like glycosylation).
2– signal processing functions are not very adaptable, they do not see pattern well, they ignore some peaks and find others in a way that is inflexible

3– N term peak height in the dodecamers and higher multimers shows variation in N term attachments.

Verge of a Dream: Friends for Life

That red brick grammar
School is no longer there.
A lefty, steve hit a soft
Ball firmly off its
Upper level wall.
Only knowing him
slightly, thought he’d
make something of
himself with that
hard high drive that
went farther (it was not
further.., there’s the
grammar learned at
Levitt avenue school). I
thought he’d do
something in life though
I only knew him lightly.
Not even the house he
was raised in
the small town pretending
To be of Scottish descent
I guess
I have no sense of what
success is…or whether
it is sweetly fragrant
though a bust
of burt Lancaster in
a hall of fame would
be titled, my friends
from childhood are
friends for life.
I know that hitting
A ball apparently
occasionally hit well
seems to be a substitute
for success as memory
substitutes for kids
I didn’t know well
enough to still be friends.