Snake Venom Storage- Fresh>Commercial

Snake Venom Instability – Willemse & Hattingh, 1978

• Abstract: Compared venom samples of rinkals (H. haemachatus), Egyptian cobra (N. haje haje), and puff adder (B arietans) via electrophoresis. Found a considerable difference between fresh and commercial samples even within species, probably because of storage conditions.
• Introduction: A lot of studies have shown that venom can be quite unstable especially when kept in liquid form. Futhermore, dessicated or dried samples lose potency over time and some studies have used samples that are as old as 26 years. Thus the differences seen between venom samples that are chalked up to geographic region, or experimental technique are actually influenced by method of storage, which is rarely taken into account. This study observes the effects of storage on the electrophoretic pattern of 3 venomous South African snake.
• Methods:
  Venom: freeze-dried and dessicated commercial samples were obtained for each of the species and electrophoresis was conducted within 24hrs of procurement. For the fresh venom, the snakes were milked using the physical restraining method and were pooled according to species (maybe we should avoid this, some papers show high variability between individuals, would we lose important proteins because they are being shown at low amounts and then getting overpowered by accumulative common bands? -> separate individual samples??? cause even over time there could be a diff? overkill??). Some of this was immediately analyzed, some were freeze-dried first and then analyzed, while others were freeze-dried and then left for 14 days. check text for more specifics
  Electrophoresis: “Electrophoresis was done on 7,5% (mlv) polyacrylamide gels in O,05M Tris-glycine buffer at pH 8,5 for 45 minutes at 160V, according to the method described byDavis (1964). This technique has the advantage of considerable resolving power while requiring minimal amounts of venom. The gels were stained with saturated Amido Black in. 7 ,5% glacial acetic acid and after destaining in 7,5% glacial acetic acid the gels were scanned in a Beckman R 110 Microzone Densitometer equipped with an integrator. The protein fractions were number” -> taken from text word for word cause i have no idea how this shit is done -_-, talked to a friend who said that electrophoresis is used to separate a compound by specific proteins according to molecular weight, SDS is used to get rid of charge.
• Results:
  Puffadders: commercial samples were compared with fresh samples that were from the same geographic region, smart cause it cuts out geographic variation. Found that both samples were pretty similar except Fraction SA16 was present in fresh and not in commercial. Also fresh venom had 90% at anode while commercial had 99%. I wonder if hemolyph or stress related markers are heavily seen in these samples given the methodology.
  Rinkals: Similar story but fresh venom had the cathode at 33% while commercial had less than 30%. Different protein bands were higher in the fresh venom and similarly for the commercial, some were also absent or almost non-existent in one and not the other. Some found solely in commercial are RAsc, RC2a, RC2b, RCa, RC,a, RCI2a.
  Egyptian cobra: basically shows that freeze-dried sample best corresponds with the fresh venom, however 2 of the bands have split into 2. also big takeaway is that different venoms from different species changed at different rates, and the Egyptian cobra was slower than rinkals.
• Discussion: given how much venom seems to change with storage duration and storage type, it is recommended to use fresh venom or fresh freeze dried venom because that antivenom will be effective against fresh venom. Commercial ones won’t be as effective because of the additional bands.